College of American Pathologists-Microbiology Checklist

Master
Microbiology Checklist
CAP Accreditation Program
College of American Pathologists

325 Waukegan Road
Northfield, IL 60093-2750
www.cap.org 08.21.2017
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Microbiology Checklist 08.21.2017
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The Checklists used for inspection by the College of American Pathologists' Accreditation Programs
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action to protect these copyrights.
All Checklists are ©2017. College of American Pathologists. All rights reserved.
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TABLE OF CONTENTS
SUMMARY OF CHANGES....................................................................................................................5
INTRODUCTION.................................................................................................................................... 8
GENERAL MICROBIOLOGY................................................................................................................ 8
PROFICIENCY TESTING.................................................................................................................................................... 8
QUALITY MANAGEMENT AND QUALITY CONTROL....................................................................................................... 9
WAIVED TESTS............................................................................................................................................................ 9
GENERAL ISSUES..................................................................................................................................................... 10
SPECIMEN COLLECTION AND HANDLING............................................................................................................. 13
REAGENTS - GENERAL............................................................................................................................................ 15
REPORTING OF RESULTS........................................................................................................................................16
INSTRUMENTS AND EQUIPMENT............................................................................................................................16
MATRIX-ASSISTED LASER DESORPTION IONIZATION TIME-OF-FLIGHT (MALDI-TOF) MASS
SPECTROMETRY..............................................................................................................................................................17
BIOSAFETY....................................................................................................................................................................... 19
BACTERIOLOGY................................................................................................................................. 22
MEDIA................................................................................................................................................................................ 22
STAINS...............................................................................................................................................................................24
REAGENTS........................................................................................................................................................................ 25
BACTERIOLOGY SUSCEPTIBILITY TESTING................................................................................................................ 28
PROCEDURES AND TESTS............................................................................................................................................ 31
RESPIRATORY SPECIMENS..................................................................................................................................... 32
URINE SPECIMENS................................................................................................................................................... 33
GENITAL SPECIMENS............................................................................................................................................... 33
STOOL SPECIMENS.................................................................................................................................................. 34
CEREBROSPINAL & OTHER BODY FLUID SPECIMENS........................................................................................35
BLOOD CULTURES.................................................................................................................................................... 36
ANAEROBIC CULTURES........................................................................................................................................... 38
WOUND SPECIMENS.................................................................................................................................................39
GAS CHROMATOGRAPHY (GC) FOR MICROBIAL IDENTIFICATION.................................................................... 39
LABORATORY SAFETY....................................................................................................................................................41
MYCOBACTERIOLOGY...................................................................................................................... 42
QUALITY CONTROL......................................................................................................................................................... 42
SPECIMEN HANDLING...............................................................................................................................................42
REPORTING OF RESULTS........................................................................................................................................43
MEDIA.......................................................................................................................................................................... 43
CONTROLS AND STANDARDS.................................................................................................................................45
RAPID METHODS....................................................................................................................................................... 46
CONCENTRATION, INOCULATION, INCUBATION...................................................................................................47
CULTURES.................................................................................................................................................................. 47
DIFFERENTIAL BIOCHEMICAL PROCEDURES....................................................................................................... 48
High Performance Liquid Chromatography (HPLC) for Microbial Identification....................................................49
MYCOLOGY......................................................................................................................................... 53
QUALITY CONTROL......................................................................................................................................................... 53
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MEDIA.......................................................................................................................................................................... 54
CONTROLS AND STANDARDS:................................................................................................................................55
MYCOLOGY SUSCEPTIBILITY TESTING........................................................................................................................58
PARASITOLOGY................................................................................................................................. 62
QUALITY CONTROL......................................................................................................................................................... 62
REAGENTS................................................................................................................................................................. 63
INSTRUMENTS AND EQUIPMENT............................................................................................................................63
STOOLS FOR OVA AND PARASITES...................................................................................................................... 64
BLOOD FILMS FOR MALARIA AND OTHER PARASITES....................................................................................... 65
VIROLOGY........................................................................................................................................... 68
QUALITY CONTROL......................................................................................................................................................... 68
REAGENTS................................................................................................................................................................. 68
CONTROLS AND STANDARDS.................................................................................................................................72
TESTS AND PROCEDURES............................................................................................................................................ 73
MOLECULAR MICROBIOLOGY......................................................................................................... 76
QUALITY MANAGEMENT................................................................................................................................................. 76
SPECIMEN HANDLING & PROCESSING........................................................................................................................ 77
ASSAY VALIDATION AND VERIFICATION......................................................................................................................78
QUANTITATIVE ASSAYS: CALIBRATION & STANDARDS.............................................................................................81
QUALITY CONTROL......................................................................................................................................................... 85
REAGENTS ...................................................................................................................................................................... 89
INSTRUMENTS..................................................................................................................................................................90
PROCEDURES & TESTS..................................................................................................................................................91
ELECTROPHORESIS..................................................................................................................................................93
MICROBIAL IN SITU HYBRIDIZATION (ISH)............................................................................................................ 94
SEQUENCING............................................................................................................................................................. 95
RESULTS REPORTING.................................................................................................................................................... 96
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ON-LINE CHECKLIST AVAILABILITY

Participants of the CAP accreditation programs may download the checklists from the CAP website
(www.cap.org) by logging into e-LAB Solutions. They are available in different checklist types and formatting
options, including:
● Master — contains ALL of the requirements and instructions available in PDF, Word/XML or Excel
formats
● Custom — customized based on the laboratory's activity (test) menu; available in PDF, Word/XML or
Excel formats
● Changes Only — contains only those requirements with significant changes since the previous checklist
edition in a track changes format to show the differences; in PDF version only. Requirements that have
been moved or merged appear in a table at the end of the file.
SUMMARY OF CHECKLIST EDITION CHANGES

08/21/2017 Edition
The information below includes a listing of checklist requirements with significant changes in the current edition
and previous edition of this checklist. The list is separated into three categories:
1. New
2. Revised:
● Modifications that may require a change in policy, procedure, or process for continued
compliance; or
● A change to the Phase
3. Deleted/Moved/Merged:
● Deleted
● Moved — Relocation of a requirement into a different checklist (requirements that have been
resequenced within the same checklist are not listed)
● Merged — The combining of similar requirements
NOTE: The listing of requirements below is from the Master version of the checklist. The customized checklist
version created for on-site inspections and self-evaluations may not list all of these requirements.
NEW Checklist Requirements
Requirement Effective Date

MIC.00375 08/17/2016
MIC.11395 08/17/2016
MIC.22675 08/17/2016
MIC.61060 08/21/2017
REVISED Checklist Requirements

MIC.11025 08/17/2016
MIC.16605 08/17/2016
MIC.18976 08/17/2016
MIC.21240 08/17/2016
MIC.21813 08/17/2016
MIC.21910 08/17/2016
MIC.21943 08/21/2017
MIC.22140 08/17/2016
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MIC.22610 08/17/2016
MIC.22640 08/17/2016
MIC.22700 08/17/2016
MIC.22830 08/17/2016
MIC.32100 08/17/2016
MIC.43100 08/17/2016
MIC.53050 08/21/2017
MIC.61140 08/21/2017
MIC.61210 08/17/2016
MIC.61310 08/17/2016
MIC.61370 08/21/2017
MIC.63220 08/17/2016
MIC.64760 08/21/2017
MIC.64770 08/21/2017
MIC.64960 08/21/2017
MIC.64968 08/21/2017
MIC.65120 08/21/2017
MIC.65130 08/21/2017
MIC.65140 08/21/2017
MIC.65200 08/17/2016
MIC.65320 08/17/2016
MIC.65330 08/21/2017
MIC.65340 08/21/2017
MIC.65550 08/21/2017
MIC.66120 08/21/2017
DELETED/MOVED/MERGED Checklist Requirements

MIC.16275 08/20/2017
MIC.17000 08/20/2017
MIC.17050 08/20/2017
MIC.21840 08/16/2016
MIC.22710 08/20/2017
MIC.31660 08/16/2016
MIC.42450 08/16/2016
MIC.43150 08/16/2016
MIC.61330 08/20/2017
MIC.63282 08/16/2016
MIC.64631 08/16/2016
MIC.64730 08/20/2017
MIC.64810 08/20/2017
MIC.64815 08/20/2017
MIC.64840 08/20/2017
MIC.64860 08/20/2017
MIC.64880 08/20/2017
MIC.64884 08/20/2017
MIC.64886 08/20/2017
MIC.64920 08/20/2017
MIC.64926 08/20/2017
MIC.64938 08/20/2017
MIC.64952 08/20/2017
MIC.64964 08/20/2017
MIC.64972 08/16/2016
MIC.64976 08/16/2016
MIC.64980 08/20/2017
MIC.64984 08/16/2016
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MIC.65000 08/20/2017
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INTRODUCTION
This checklist is used in conjunction with the All Common and Laboratory General Checklists to inspect a
microbiology laboratory section or department.
Certain requirements are different for waived versus nonwaived tests. Refer to the checklist headings and
explanatory text to determine applicability based on test complexity. The current list of tests waived under CLIA
may be found at http://www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfClia/analyteswaived.cfm.
Note for non-US laboratories: Checklist requirements apply to all laboratories unless a specific disclaimer of
exclusion is stated in the checklist.
GENERAL MICROBIOLOGY
Requirements in this section apply to ALL of the subsections in the microbiology laboratory (bacteriology,
mycobacteriology, mycology, parasitology, molecular microbiology, and virology).
PROFICIENCY TESTING
Inspector Instructions:
● Are proficiency testing samples tested to the same level as clinical specimens?
● Select a representative clinical report of each culture type. Compare the extent of
reporting for the relevant proficiency testing sample.
MIC.00350 PT Extent of Testing Phase II

Organisms in proficiency testing specimens are identified to the same level as those from
patient samples.
NOTE: If the laboratory's proficiency testing reports include incomplete identifications (e.g. "Gram
positive cocci" or "Mycobacterium species, not tuberculosis"), it must indicate that this matches
the information produced by the laboratory's internal capabilities in patient reports. In other
words, patient reports cannot be more specific than the identification level reporting in proficiency
testing, unless the former contain more specific information provided by referral laboratories.
**NEW** 08/17/2016
MIC.00375 PT for Susceptibility Testing Phase II
If any susceptibility testing is performed on-site, the laboratory participates in a
proficiency testing program for the related subspecialty (e.g. bacteriology, mycology).
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Evidence of Compliance:
✓ Records of proficiency testing performance
QUALITY MANAGEMENT AND QUALITY CONTROL

● Sampling of QC policies and procedures
● Sampling of QC records
● Sampling of employee records of morphologic observation correlation
● How do you determine when QC is unacceptable and when corrective actions are
needed?
● How do you ensure consistency among personnel performing microscopic
morphology?
● Select several occurrences in which QC is out of range and follow records to

determine if the steps taken follow the laboratory procedure for corrective action
● Use QC data to identify tests that utilize internal quality control processes and confirm
that the tests have an individualized quality control plan (IQCP) approved by the
laboratory director, when required
WAIVED TESTS
MIC.10060 QC - Waived Tests Phase II

The laboratory follows manufacturer's instructions for quality control, records and
reviews results for acceptability prior to reporting patient results.
NOTE: Quality control must be performed according to manufacturer's instructions. Testing

personnel or supervisory staff must review quality control data on days when controls are run
prior to reporting patient results. The laboratory director or designee must review QC data at
least monthly or more frequently if specified in the laboratory QC policy.
With respect to internal controls, acceptable control results must be recorded, at a minimum,
once per day of patient testing for each device.*
*Acceptable internal control results need not be recorded, if (and only if) an unacceptable
instrument control automatically locks the instrument and prevents release of patient results.
✓ Written procedure consistent with manufacturer's instructions for each waived test AND
✓ Records showing confirmation of acceptable QC results
MIC.10070 QC Corrective Action - Waived Tests Phase II

There are records of corrective action when quality control results exceed the acceptable
range.
NOTE: The remaining requirements in this checklist on quality control do not apply to waived tests.
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GENERAL ISSUES
MIC.11015 QC Handling Phase II

Control specimens are tested in the same manner and by the same personnel as patient
samples.
NOTE: QC specimens must be analyzed by personnel who routinely perform patient testing. This
does not imply that each operator must perform QC daily, so long as each instrument and/or test
system has QC performed at required frequencies, and all analysts participate in QC on a regular
basis. To the extent possible, all steps of the testing process must be controlled, recognizing that
preanalytic and postanalytic variables may differ from those encountered with patients.
✓ Records reflecting that QC is run by the same personnel performing patient testing
REFERENCES
1) Department of Health and Human Services, Centers for Medicare and Medicaid Services. Clinical laboratory improvement
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7166 [42CFR493.1256(d)(8)].1256(f)]
MIC.11016 Commercial Product - QC Phase II

When using a commercial product, QC is performed according to the manufacturer's
instructions or CAP Checklist requirements, whichever is more stringent.
NOTE: This includes, but is not limited to, antimicrobial susceptibility testing/identification (AST/
ID) systems.
MIC.11017 QC Confirmation of Acceptability Phase II

Control results are reviewed for acceptability before reporting patient results.
✓ Written policy stating that controls are reviewed and acceptable prior to reporting patient
results AND
✓ Evidence of corrective action taken when QC results are not acceptable
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24)1992(Feb 28):7166 [42CFR493.1256(f)]
MIC.11018 QC Corrective Action Phase II

There are records of corrective action when control results exceed defined acceptability
limits.
NOTE: Patient/client test results obtained in an analytically unacceptable test run or since the last
acceptable test run must be re-evaluated to determine if there is a significant clinical difference
in patient/client results. Re-evaluation may or may not include re-testing patient samples,
depending on the circumstances.
Even if patient samples are no longer available, test results can be re-evaluated to search for
evidence of an out-of-control condition that might have affected patient results.
The corrective action for tests that have an Individualized Quality Control Plan (IQCP) approved
by the laboratory director must include an assessment of whether further evaluation of the risk
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assessment and quality control plan is needed based on the problems identified (e.g. trending for
repeat failures, etc.).
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Oct 1):1046[42CFR493.1282(b)(2)]
MIC.11020 Monthly QC Review Phase II

Quality control data are reviewed and assessed at least monthly by the laboratory director
or designee.
NOTE: The review of quality control data must be recorded and include follow-up for outliers,
trends, or omissions that were not previously addressed.
The QC data for tests performed less frequently than once per month should be reviewed when
the tests are performed.
The review of quality control data for tests that have an IQCP approved by the laboratory director
must include an assessment of whether further evaluation of the risk assessment and quality
control plan is needed based on problems identified (e.g. trending for repeat failures, etc.).
✓ Records of QC review including follow-up for outliers, trends or omissions
**REVISED** 08/17/2016
MIC.11025 Alternative Control Procedures Phase II
If the laboratory performs test procedures for which control materials are not
commercially available, there are written procedures for an alternative mechanism to
detect immediate errors and monitor test system performance over time. The performance
of alternative control procedures must be recorded.
NOTE: "Performance" includes elements of accuracy, precision, and clinical discriminating

power. Examples of alternative procedures may include split sample testing with another method
or with another laboratory, the testing of previously tested patient specimens in duplicate, testing
of patient specimens in duplicate, or other defined processes approved by the laboratory director.
✓ Written procedures for alternative quality control AND
✓ Records of alternative control procedures
REFERENCES
1) Elder BL, et al. Verification and Validation of Procedures in the Clinical Microbiology Laboratory. Cumitech 31, February 1997. ASM
Press; Washington DC
2) Sharp S and Clark R. Verification and Validation of Procedures in the Clinical Microbiology Laboratory. Cumitech 31a. 2009. ASM
Press: Washington, DC
amendments of 1988; final rule. Fed Register. 2003(Jan 24): [42CFR493.1256(h)].
MIC.11060 Culture Result Reporting Phase I

If the laboratory is reporting culture results other than simply "growth/no growth," the
laboratory has the ability to perform Gram stains as part of its bacterial identification
process.
NOTE: The performance of a Gram stain on colonies from a culture plate may be a necessary
procedure for guiding culture workup and in confirming the identification of organisms, especially
when atypical findings are noted during the workup.
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Personnel performing Gram stains for this purpose are subject to competency assessment.
Requirements for proficiency testing must be met through participation in the bacterial culture
proficiency testing programs.
MIC.11350 Morphologic Observation Assessment Phase II

The microbiology laboratory at least annually assesses morphologic observations among
personnel performing Gram, trichrome and other organism stains, to ensure consistency.
NOTE: Suggested methods to accomplish this include:
1. Circulation of organisms with defined staining characteristics, and/or

2. Multi-headed microscopy, and/or
3. Use of photomicrographs with referee and participant identifications (e.g. former CAP
microbiology Surveys or other photomicrographs from teaching collections)
4. Use of digital images
✓ Written procedure defining the method(s) and criteria used for evaluation of consistency AND
✓ Employee records of morphology assessment
REFERENCES
1) Flournoy DJ. Interpreting the sputum gram stain report. Lab Med. 1998;29:763-768
MIC.11375 Taxonomy Changes Phase I

The laboratory incorporates taxonomic changes that potentially affect the choice of
appropriate antimicrobials to report and/or the interpretative breakpoints to use.
NOTE: The genus and/or species names of microorganisms may change as new methods
are applied to their taxonomy. This can impact the antimicrobials that should be reported for
that organism. It may also impact which breakpoints are used for reporting. For example,
Actinobacillus actinomycetemcomitans was moved to the genus Haemophilus in 1985 and then
to the new genus Aggregatibacter in 2006. The antimicrobials differ for Haemophilus species
(CLSI M100, Table 2E) versus Aggregatibacter species (CLSI M45, Table 7). The laboratory
should have a policy ensuring that clinically relevant taxonomic changes are incorporated into
reporting patient and proficiency testing results even when commercial identification systems
have not been updated.
Taxonomic nomenclature is not valid until published in the International Journal of Systematic
& Evolutionary Microbiology (IJSEM). For laboratories participating in the CAP's proficiency
programs for microbiology, the Participant Summary Report Final Critique is a good source of
information as the Microbiology Resource Committee provides periodic updates in taxonomy
through educational challenges.
Additional information for specific specialties may be found on-line using web sites, such as the
following:
For bacteriology
● http://www.bacterio.net/-classification.html
● http://www.dsmz.de/bacterial-diversity/prokaryotic-nomenclature-up-to-date.html
● http://www.zipcodezoo.com
● http://enews.patricbrc.org/
For mycology:
● http://mycobank.org
● http://www.mycology.adelaide.edu.au
● http://www.fungaltaxonomy.org
For parasitology:
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● http://www.cdc.gov/dpdx/
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Methods for Antimicrobial Dilution and Disk Susceptibility Testing of Infrequently
Isolated or Fastidious Bacteria; Approved Guideline; 2nd ed. CLSI document M45-ED3. Clinical and Laboratory Standards Institute,
Wayne, PA, 2016.
2) Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing; 27th ed. CLSI
supplement M100-S27. Clinical and Laboratory Standards Institute, Wayne, PA, 2017.
**NEW** 08/17/2016
MIC.11395 Referral of Isolates for Susceptibility Testing Phase I
If the laboratory is unable to perform susceptibility testing on-site, there is a mechanism
to refer clinically significant isolates for which susceptibility testing is deemed necessary
(e.g. isolates obtained from blood or other sterile sites).
✓ Written policy defining situations where isolates must be referred for susceptibility testing
AND
✓ Records of referral of isolates for susceptibility testing
SPECIMEN COLLECTION AND HANDLING
Culture specimens are often collected by nurses or others outside the laboratory. An important aspect of quality
control is the provision of adequate instructions to ensure proper collection and handling of specimens before
they are received by the laboratory.
● Sampling of specimen collection and handling policies and procedures
● Sampling of requisitions for completeness
● Sampling of microbiology specimens (transport media, timely delivery, labeling)
● What is your course of action when you receive unacceptable microbiology

specimens?
MIC.13175 Viral Culture Specimens Phase I

Specimens for viral culture are collected appropriately and transported to the laboratory
without delay.
NOTE: The laboratory must provide procedures for the appropriate collection, transport and
storage of all specimen types tested in the laboratory. Specimens should be delivered to the
laboratory promptly, ideally within 2-4 hrs of sample collection and preferably within 1 day
of collection. This may not be possible for laboratories that refer samples to offsite referral
laboratories for viral testing. In these instances samples must be stored and shipped under
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conditions that would preserve the integrity of the sample. Unless otherwise indicated, specimens
should be refrigerated or frozen depending on the duration of storage prior to testing.
REFERENCES
1) Clinical and Laboratory Standards Institute. Viral Culture; Approved Guideline. CLSI document M41-A. Clinical and Laboratory
Standards Institute, Wayne, PA, 2006.
2) Ginocchio, CC. Quality Assurance in Clinical Virology. In: Spector S, Hodinka RL, Young SA, editors. Clinical Virology Manual.
Fourth Edition. Washington: ASM Press; 2009.p. 3-17
MIC.13200 Requisitions Phase I

Requests for analysis include source of specimen, test or tests requested and, when
appropriate, type of infection and/or organism expected.
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7162 [42CFR493.1241(c)]
MIC.13250 Specimen Collection/Handling Phase II

There are written instructions for microbiology specimen collection and handling that
include all of the following.
1. Method for proper collection of culture specimens from different sources
2. Proper labeling of culture specimens
3. Use of appropriate transport media when necessary
4. Policies for safe handling of specimens (tightly sealed containers, no external
spillage)
5. Need for prompt delivery of specimens to ensure minimum delay and
processing (e.g. CSF, wound cultures, anaerobes)
6. Method for preservation of specimens if processing is delayed (e.g.
refrigeration of urines)
NOTE: Manufacturer's recommendations must be followed when there is a delay in delivery or

processing of specimens for automated instruments (e.g. blood culture instruments).
REFERENCES
1) Jorgensen JH (ed), et al. Manual of Clinical Microbiology. 11th ed. Washington, DC: ASM Press; 2015.
MIC.13275 Specimens for Molecular Amplification Phase II

The laboratory has written procedures for the handling of specimens that will be tested
using molecular amplification methods.
NOTE: Special precautions must be taken to avoid sample cross-contamination that may not
affect culture-based methods but may lead to false positive results when tested using molecular
amplification methods. For example, proper methods to prevent cross-contamination must be
used when samples are processed in the same biohazard hood in which virus cultures are
manipulated post-inoculation. Please refer to the Molecular Microbiology section of this checklist.
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Collection, Transport, Preparations, and Storage of Specimens for Molecular
Methods; Approved Guideline. CLSI document MM13-A (ISBN 1-56238-591-7). Clinical and Laboratory Standards Institute, 940
West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2005.
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REAGENTS - GENERAL
● Sampling of test procedures for QC
● Sampling of reagent QC records
Additional requirements are in the REAGENTS section of the All Common Checklist.
The following generic requirements apply to all subsections of the Microbiology Laboratory for nonwaived testing
only.
MIC.14583 Direct Antigen Test QC Phase II

For nonwaived direct antigen tests on patient specimens, positive and negative controls
are tested and recorded at least daily, or more frequently if specified in the manufacturer's
instructions, laboratory procedure, or CAP Checklist.
NOTE: This requirement pertains to nonwaived tests with a protein, enzyme, or toxin which
acts as an antigen. Examples include, but are not limited to: Group A Streptococcus antigen, C.
difficile toxin, fecal lactoferrin and immunochemical occult blood tests. For panels or batteries,
controls must be employed for each antigen sought in patient specimens.
If an internal quality control process (e.g. electronic/procedural/built-in) is used instead of an
external control material to meet daily quality control requirements, the laboratory must have an
individualized quality control plan (IQCP) approved by the laboratory director to address the use
of the alternative control system. Please refer to the individualized Quality Control Plan section of
the All Common Checklist for the eligibility of tests for IQCP and requirements for implementation
and ongoing monitoring of an IQCP.
For each test system that requires an antigen extraction phase, as defined by the manufacturer,
the system must be checked with an appropriate positive control that will detect problems in
the extraction process. If an IQCP is implemented for the test, the laboratory's quality control
plan must define how the extraction phase will be monitored, as applicable, based on the risk
assessment performed by the laboratory and the manufacturer's instructions.
✓ Written QC procedures AND
✓ Records of QC results including external and electronic/procedural/built-in control systems
AND
✓ Manufacturer's product insert or manual
REFERENCES
amendments of 1988, final rule. Fed Register. 2003(Jan 24): [42CFR493.1261(a)]
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REPORTING OF RESULTS
● Sampling of patient preliminary reports
MIC.15000 Preliminary Reports Phase I

When indicated, preliminary reports are promptly generated.
✓ Written policies defining when preliminary results are issued
MIC.15020 Azoospermic Specimen Result Reporting Phase I

For azoospermic and post-vasectomy seminal fluid specimens, the laboratory clearly
communicates the findings of the assay and either employs a concentrating technique on
seminal fluid or includes a comment in the patient report indicating that a concentrating
technique was not performed.
NOTE: Without a concentration technique, the presence of both motile and non-motile sperm
may not be detected. The method for detection of motile and non-motile sperm and the
laboratory findings must be clearly communicated on the patient report so that the clinician can
interpret the results in context to the method performed. The decision on the method used and
extent of testing to be performed should be made in consultation with the medical staff served.
The American Urological Association (AUA) Vasectomy Guideline recommends a careful
evaluation of an uncentrifuged specimen, and does not recommend centrifugation of the
specimen for further assessment. The AUA Guideline also recommends reporting both the
presence and absence of sperm and presence or absence of sperm motility on the patient report.
If no sperm are seen in the uncentrifuged specimen, the guideline recommends reporting that the
presence of sperm is below the limit of detection.
✓ Patient report with concentration findings or appropriate comment indicating that
concentration was not performed
REFERENCES
1) Evaluation of the Azoospermic Male. Fertil Steril. 2008; 90 (S74-7)
2) Diagnostic Evaluation of the Infertile Male: A Committee Opinion. Fertil Steril. 2012; 98:294-301
3) American Urological Association (AUA) Guideline. American Urological Association Education and Research, Inc. 2012; amended
2015. https://www.auanet.org/guidelines/vasectomy-(2012-amended-2015)
4) Vasectomy Update 2010. Can Urol Assoc J. 2010 October; 4(5):306-309
INSTRUMENTS AND EQUIPMENT
The checklist requirements in this section should be used in conjunction with the requirements in the All
Common Checklist relating to instruments and equipment.
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● Sampling of pipette/diluter checks
● Incubators (adequate space, maintained)
MIC.16150 Pipettors and Diluters Phase II

Pipettes, microtiter diluters or automatic dispensers that are used for quantitative
dispensing of material are checked for accuracy and reproducibility at defined intervals
(at least annually), with results recorded.
NOTE: Pipette checks must be performed following manufacturer's instructions, at minimum, and
as defined in laboratory procedure.
This requirement is not applicable for precalibrated inoculation loops that are used in the direct
plating of clinical specimens such as urine cultures.
✓ Written procedure detailing method for checking the accuracy and reproducibility of pipettes
REFERENCES
1) Curtis RH. Performance verification of manual action pipets. Part I. Am Clin Lab. 1994;12(7):8-9
2) Curtis RH. Performance verification of manual action pipets. Part II. Am Clin Lab. 1994;12(9):16-17
3) Perrier S, et al. Micro-pipette calibration using a ratiometric photometer-reagent system as compared to the gravimetric method. Clin
Chem. 1995;41:S183
4) Clinical and Laboratory Standards Institute. Laboratory Instrument Implementation, Verification, and Maintenance; Approved
Guideline. CLSI Document GP31-A. Clinical and Laboratory Standards Institute, Wayne, PA; 2009.
5) Johnson B. Calibration to dye for: Artel's new pipette calibration system. Scientist. 1999;13(12):14
6) Connors M, Curtis R. Pipetting error: a real problem with a simple solution. Parts I and II. Am Lab News. 1999;31(13):20-22
7) Skeen GA, Ashwood ER. Using spectrophotometry to evaluate volumetric devices. Lab Med. 2000;31:478-479
MIC.16550 Adequate Incubators Phase I

There are sufficient, clean, and well-maintained incubators available at specified
temperature ranges.
MATRIX-ASSISTED LASER DESORPTION IONIZATION

TIME-OF-FLIGHT (MALDI-TOF) MASS SPECTROMETRY
This section applies to laboratories using MALDI-TOF systems to perform organism identification. Refer to the
Test Method Validation section in the All Common Checklist for validation requirements pertinent to laboratory-
developed tests.
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● Sampling of mass spectrometer policies and procedures
● Identification criteria compliance
● Sampling of calibration and control records
● How does your laboratory verify assay performance each day of use?
MIC.16575 Instrument Operation Phase II

There are written procedures for the operation and calibration of the mass spectrometer.
MIC.16595 Mass Spectrometer Calibration Phase II

A calibration control is run each day of patient testing, with each change in target plate, or
according to manufacturer's recommendations and these records are maintained.
NOTE: Acceptable tolerance limits for calibration parameters must be defined, and records
maintained.
✓ Records of calibration
**REVISED** 08/17/2016
MIC.16605 Mass Spectrometer Controls Phase II
Appropriate control organisms or calibrators are tested on each day of patient testing.
NOTE: Appropriate controls would include at least one bacterium, with a representative yeast
and mycobacterium also being run if these organisms are being tested for that day/routinely. For
FDA-approved platforms, the organisms or calibrator(s) required by the manufacturer must be
used.
For laboratory developed tests, choice of control organisms is at the Laboratory Director's
discretion. Control organisms must be subjected to the same testing conditions throughout the
testing procedure as patient specimens. An extraction control should be included if any of the
organisms being tested are run with extraction.
In formats of testing where a target is reused, a blank control needs to be run after each
cleansing to assess the cleanliness of the target (demonstrating a lack of peaks prior to testing).
✓ Written policy defining QC requirements AND
✓ QC records at defined frequency
MIC.16615 Mass Spectrometer Reagent Grade Phase II

Reagents and solvents are of appropriate grade.
NOTE: Only the manufacturer's specified grade of solvents are used for this procedure. This may
be HPLC-grade or other reagent grades as indicated.
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✓ Reagent logs
MIC.16625 Mass Spectrometer Consumables Phase II

Consumables appropriate to the instrument and assay are required.
NOTE: For FDA-approved platforms, consumables utilized may be specified by the

manufacturer. Deviation from the manufacturer's recommendation must be validated.
✓ Consumable logs AND
✓ Validation of alternative consumables not specified by the manufacturer
BIOSAFETY
Items in this section apply to ALL areas of the microbiology laboratory. Additional items for specific subsections
(bacteriology, mycobacteriology, mycology, parasitology, and virology) are found under the Laboratory Safety
subsections for each of those areas.
● Sampling of biosafety policies and procedures
● Sampling of bench top decontamination logs
● Records of biological safety cabinet certification
● How would you recognize a potential agent of bioterrorism? What action would you
take if you encountered a suspect organism?
MIC.18968 Agents of Bioterrorism Phase II

The microbiology laboratory has policies and procedures for the recognition and safe
handling of isolates that may be used as agents of bioterrorism.
NOTE: Microorganisms likely to be utilized as biological weapons include Bacillus anthracis

(anthrax), Brucella species (brucellosis), Clostridium botulinum (botulism), Francisella tularensis
(tularemia), Yersinia pestis (plague) and variola major (smallpox).
As part of an institution-wide plan to prepare and respond to a bioterrorism event, the
microbiology laboratory should have policies and procedures for the recognition of isolates that
may be used as agents of bioterrorism.
Safe handling includes such activities as handling organisms under a certified biological safety
cabinet, and not subjecting the isolates to identification utilizing automated instruments.
REFERENCES
1) Snyder JW. Role of the hospital-based microbiology laboratory in preparation and response to a bioterrorism event. J Clin Microbiol.
January, 2003
2) Gilchrist MJR. Laboratory Safety, Management, and Diagnosis of Biological Agents Associated with Bioterrorism
3) Robinson-Dunn B. The microbiology laboratory's role in response to bioterrorism. Arch PatholLab Med. March 2002; 126
4) Morse SA. Bioterrorism: Laboratory Security. Lab Med. June 2001
5) Sewell, DL. Laboratory safety practices associated with potential agents of biocrime or bioterrorism. J. Clin. Microbiology. July
2003;41(7):2801-2809
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6) https://www.asm.org/index.php/guidelines/sentinel-guidelines
**REVISED** 08/17/2016
MIC.18976 Bioterrorism Response Plan Phase I
The laboratory is recognized in the institution's bioterrorism response plan and the role of
the laboratory is outlined in the plan.
✓ Organizational bioterrorism plan describing the role of the laboratory
REFERENCES
1) Snyder JW. Role of the hospital-based microbiology laboratory in preparation and response to a bioterrorism event. J Clin Microbiol.
January, 2003
2) Gilchrist MJR. Laboratory Safety, Management, and Diagram of Biological Agents Associated with Bioterrorism
3) Robinson-Dunn B. The microbiology laboratory's role in response to bioterrorism. Arch PatholLab Med. March 2002; 126
4) Morse SA. Bioterrorism: Laboratory Security. Lab Med. June 2001
MIC.18985 Spill Handling Phase II

There are written policies and procedures for handling spills of contaminated materials.
REFERENCES
1) Department of Health and Human Services. Biosafety in Microbiological and Biomedical Laboratories, 5th ed. Washington, DC: HHS
Publishing No. (CDC) 21-1112, December 2009
MIC.19010 Bench Top Decontamination Phase II

There are records of daily decontamination of bench tops.
MIC.19035 Safe Specimen Processing Phase II

There are written policies and procedures for the safe handling and processing of
specimens.
NOTE: Suggested topics to be considered in the policies and procedures for the safe handling
and processing of specimens include the need for tight sealing of containers, avoiding spills
of hazardous materials, requirements for wearing gloves, the need for respirator protection,
availability and use of vaccinations, and the potential hazards of sniffing plates.
REFERENCES
1) Jamison R, et al. Laboratory Safety in Clinical Microbiology, Cumitech 29, July 1996, ASM Press; Washington DC
2) Fleming DO, Hunt DL. Biological Safety, Principles and Practices, 3rd ed. ASM Press; Washington DC
MIC.19060 Biosafety Levels Phase II

Policies and procedures have been developed to minimize the occupational risk of
exposure to infectious agents handled in the microbiology laboratory, in accordance
with current recommendations regarding the biosafety levels for working with different
organisms.
NOTE: The laboratory director is responsible for the maintenance of precautions in the laboratory
to minimize the risk of personnel infection. Precautions must be appropriate for the types of
organisms tested and the nature of the studies performed.
Each level consists of combinations of equipment, processes and techniques, and laboratory
design that are appropriate for the type of laboratory and infectious agent handled.
REFERENCES
2) Richmond J. "Arthropod Borne Diseases", Anthology of Biosafety VI: American Biological Safety Association, Mundelein, IL April
2003
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MIC.19160 Biosafety Levels Phase II

Engineering and work practice controls appropriate to the Biosafety level of the laboratory
are defined and implemented.
NOTE: Each increasing BSL number (1 to 4) implies increased occupational risk from exposure
to an agent or performance of a process, and therefore is associated with more stringent control
and containment practices.
REFERENCES
2) Richmond J. BSL-4 Laboratories. Anthology of Biosafety V: American Biological Safety Association, Mundelein, IL January 2002
3) Richmond J. Biosafety Level 3. Anthology of Biosafety VII: American Biological Safety Association, Mundelein, IL December 2003
MIC.19840 Biological Safety Cabinet Phase II

A biological safety cabinet (BSC) or hood is available for handling specimens or
organisms considered highly contagious by airborne routes.
✓ Maintenance schedule of BSC function checks AND
✓ Records of testing and certification
REFERENCES

The biological safety cabinet (BSC) is certified at least annually to ensure that filters are
functioning properly and that airflow rates meet specifications.
REFERENCES
1) NSF/ANSI Standard 49-2012 Biosafety Cabinetry: Design, Construction, Performance and Field Certification. Ann Arbor, MI: NSF;
2012.
2) Kruse RH, Puckett WH, Richardson JH. Biological safety cabinetry. Clin Microbiol Rev. 1991;4(2):207-241.
3) Kimman TG, Smit E, Klein MR. Evidence-based biosafety: a review of the principles and effectiveness of microbiological containment
measures. Clin Microbiol Rev. 2008;21(3):403-425. doi: 10.1128/CMR.00014-08.
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BACTERIOLOGY
MEDIA
● Sampling of media QC policies and procedures
● Sampling of media supplier records of QC
● Sampling of records for QC performed by the laboratory
● Sampling of media (expiration date, condition, contamination)
● What is your QC procedure when receiving a new lot of media?
● Follow a shipment of new media from receipt, examination and QC (if applicable).
Determine if practice follows laboratory policy.
MIC.21220 Inspection of Media Shipments Phase I

The laboratory has records showing that each shipment of purchased media is examined
for breakage, contamination, appearance, and evidence of freezing or overheating.
REFERENCES
1) Clinical and Laboratory Standards Institute. Quality Control for Commercially Prepared Microbiological Culture Media; Approved
Standard; 3rd ed. CLSI document M22-A3. Clinical and Laboratory Standards Institute, Wayne, PA, 2004.
**REVISED** 08/17/2016
MIC.21240 Media QC - Purchased Phase II
An appropriate sample from each lot and shipment of each purchased medium is checked
before or concurrent with initial use for each of the following:
1. Sterility
2. Ability to support growth by means of stock cultures or by parallel testing with
previous lots and shipments
3. Biochemical reactivity, where appropriate
NOTE: The laboratory must have records showing that all media are sterile, able to support
growth, and are appropriately reactive biochemically. This will ordinarily require that the
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laboratory maintain a stock of reference organisms and test the media before or concurrent with
use.
An individualized quality control plan (IQCP), including all required elements of IQCP, may be
implemented by the laboratory to allow for the acceptance of the quality control performed by
the media supplier for media listed as "exempt" in the CLSI/NCCLS Standard M22-A3, Quality
Control for Commercially Prepared Microbiological Culture Media. The media supplier's records
must be maintained and show that the QC performed meets the CLSI/NCCLS standard and
checklist requirements. Please refer to the Individualized Quality Control Plan section of the All
Common Checklist for the requirements for implementation and ongoing monitoring of an IQCP.
End user quality control must be performed on the following, regardless of the exempt status:
● Campylobacter agar;
● Chocolate agar;
● Media for the selective isolation of pathogenic Neisseria;
● Other media not listed on Table 2 of M22-A3 (e.g. dermatophyte test medium);
● Media used for the isolation of parasites, viruses, Mycoplasmas, Chlamydia;
● Mueller-Hinton media used for antimicrobial susceptibility tests; or
● Media commercially prepared and packaged as a unit or system consisting of two or
more different substrates, primarily used for microbial identification.
Laboratories receiving media from media suppliers must have records showing that the quality
control activities performed by the media supplier meet the CLSI/NCCLS Standard M22-A3,
or are otherwise equivalent. The laboratory director may wish to have a signed contractual
arrangement with his/her selected media supplier to cover all expected quality control and
documentation thereof.
Laboratories using exempt media that have not implemented an IQCP or are using media that
do not qualify for an IQCP must continue to test each lot and shipment of media and maintain
records of such testing.
✓ Written procedure for QC on new lot numbers or shipments of purchased medium AND
✓ Individualized quality control plan for the media approved by the laboratory director, as
applicable AND
✓ Records of media quality control
REFERENCES
2) Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Disk Susceptibility Tests; Approved Standard.
12
th ed. CLSI document M02-A12. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
MIC.21300 Media QC - Laboratory Prepared Phase II

For microbiology media prepared by the laboratory, there are records showing that an
appropriate sample of each medium is checked before or concurrent with initial use for
each of the following:
1. Sterility
previous batches
3. Biochemical reactivity (where appropriate)
✓ Written procedure for testing media prepared by the laboratory
MIC.21420 Media Visual Examination Phase II

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All media are in visibly satisfactory condition (with expiration date, plates smooth,
adequately hydrated, uncontaminated, appropriate color and thickness, tubed media not
dried or loose from sides).
MIC.21460 Quality Control Organisms Phase II

Quality control organisms are used to check stains, reagents and susceptibility test
methods.
NOTE:
1. Quality control organisms may be ATCC strains or well characterized laboratory
strains unless specified by the manufacturer
2. Quality control organisms are maintained in a manner to preserve their bioreactivity,
phenotypic characteristics and integrity
REFERENCES
1) Jones RN, et al. Method preferences and test accuracy of antimicrobial susceptibility testing. Updates from the College of American
Pathologists microbiology surveys program (2000). Arch Pathol Lab Med. 2001;125:1285-1289
STAINS
● Sampling of staining policies and procedures
● Sampling of stain QC records/logs
MIC.21530 Direct Gram Stain Procedures Phase I

There are written policies for use of Gram stain results to provide a preliminary
identification of organisms, evaluate specimen quality when appropriate, and to guide
work-up of cultures.
NOTE: The laboratory should have policies for the interpretation of the Gram stain, including
the quantification, stain reaction, and morphotypes of organisms and cells (e.g. neutrophils or
squamous epithelial cells). The policy should address correlation of direct Gram stain results with
final culture results.
This does not mean that interpretation of the Gram stain morphology suggesting a specific
organism identification (e.g. gram positive diplococci morphologically suggestive of
pneumococcus) is required.
✓ Written policy for Gram stain (laboratories may use the correlation of Gram stain results with
the final culture results as a component of the QC program)
MIC.21540 Gram Stain QC Phase II

Quality control of Gram stain reagents is performed for intended reactivity and recorded
for each new batch of stains and at least weekly against known gram-positive and gram-
negative quality control organisms.
NOTE: Personnel who perform Gram stains infrequently must run a gram-positive and gram-
negative control each day of testing.
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✓ Written procedure for Gram stain QC
REFERENCES
1) August, Hindler, Huber, Sewell. Quality control and quality assurance practices. In: Clinical microbiology, Cumitech 3A. Washington,
DC: American Society for Microbiology, 1990
amendments of 1988, final rule. Fed Register. 2003(Jan 24):3708 [42CFR493.1261(a)(2)]
MIC.21560 Non-Immunofluorescent Stain QC Phase II

Quality control of all non-immunofluorescent, non-immunologic-based stains (other
than Gram stains) is performed and recorded with a positive and negative quality control
organism for intended reactivity each day of use, and for each new batch, lot number and
shipment.
NOTE: Refer to MIC.51160 for requirement pertaining to parasitology permanent stains.

✓ Written procedure for QC of non-immunofluorescent stains
MIC.21570 Fluorescent Stain QC Phase II

Quality control of fluorescent stains is performed for positive and negative reactivity each
time of use.
✓ Written procedure QC of fluorescent stain
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7146 [42CFR493.1256(e)(3); 493.1273(a)]
REAGENTS
● Sampling of reagent QC policies and procedures
● Sampling of reagent QC records
● CO2 monitoring procedure and CO2 recording log
● Anaerobic incubation condition monitoring records
● Campylobacter incubation condition records
● What is your QC policy when receiving a new lot of identification system materials?
MIC.21624 Reagent QC Phase II

Positive and negative controls are tested and results recorded for each new batch, lot
number, and shipment of reagents, disks/strips and stains.
NOTE: Reagents subject to this requirement include (but are not limited to) catalase, coagulase
(including latex methods), oxidase and indole reagents; bacitracin, optochin, streptococcal
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grouping reagents, ONPG, X, V, and XV disks/strips. This does not include tests for antimicrobial
susceptibility.
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24): 3708 [42CFR493.1256 (e) (1) and (2)]
MIC.21626 Identification System QC Phase II

Appropriate positive and negative control organisms are tested and results recorded for
each new lot and shipment of reagents used in bacterial identification systems.
NOTE: An individualized quality control plan (IQCP) including all required elements of IQCP, may
be implemented by the laboratory to allow for the use of streamlined QC for commercial microbial
identification systems (MIS). Refer to the individualized Quality Control Plan section of the All
Streamlined QC must be performed, as specified by the manufacturer's instructions without
modification. The laboratory may use additional QC organisms in addition to those required for
the streamlined QC. In order to qualify for streamlined QC, the user must fulfill initial and ongoing
requirements as defined by the manufacturer and CLSI Guideline M50-A, Quality Control for
Commercial Microbial Identification Systems, including the retention of test system verification
and historical QC review as long as the streamlined QC is used, but in no case for less than two
years.
For user-developed identification systems, commercial systems for which a streamlined QC
process has not been developed, or any commercial system whose use is altered in any way
from the manufacturer's instructions, all biochemical tests in each new lot number and shipment
must be evaluated with known positive and negative control organisms, to assure appropriate
reactivity.
Any test (e.g. oxidase test) required for interpretation of MIS results that is not part of the MIS
cannot be included in MIS streamlined QC procedures. QC requirements for such tests, including
the use of positive and negative controls for each new batch, lot number and shipment are given
in MIC.21624 (Reagent QC).
✓ Written procedure for QC on new lot numbers or shipments of reagents for each MIS AND
✓ Individualized quality control plan for the MIS approved by the laboratory director, as
applicable AND
✓ Records of MIS quality control
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3708 [42CFR493.1256(e)(1)]
2) Clinical and Laboratory Standards Institute (CLSI). Quality Control for Commercial Microbial Identification Systems. CLSI document
M50-A (ISBN 1-56238-675-1). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania
19087-1898 USA, 2008.
MIC.21628 Antisera QC Phase I

Positive and negative controls are tested and results recorded for each new batch, lot
number and shipment of antisera when prepared or opened and once every 6 months
thereafter (e.g. Salmonella/Shigella antisera).
REFERENCES
amendments of 1988; final rule. Fed Register. 2003 (January 24):3708 [42CFR493.1261(a)(3)]
MIC.21632 Beta-Lactamase QC Phase II

Positive and negative controls are tested and results recorded for beta-lactamase (other
than Cefinase ®) tests on each day of use.
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NOTE: Beta lactamase tests using Cefinase ® need be checked only with each batch, lot number
and shipment.
REFERENCES
amendments of 1988; final rule. Fed Register. 2003 (January 24):3708 [42CFR493.1261 (a) (1)]
MIC.21812 Anaerobic Incubation Conditions QC Phase I

There are records that anaerobic incubation systems (e.g. jars, chambers, bags) are
checked for adequate anaerobic conditions with methylene blue strips, fastidious
anaerobic organisms or other appropriate procedures.
REFERENCES
1) Clinical and Laboratory Standards Institute. Principles and Procedures for Detection of Anaerobes in Clinical Specimens; Approved
Guideline. CLSI document M56-A. Clinical and Laboratory Standards Institute, Wayne, PA; 2014
**REVISED** 08/17/2016
MIC.21813 CO2 Incubator Levels Phase I
CO2 incubators are checked daily for adequate CO2 levels, with recording of results.
NOTE: Some organisms require CO2 to grow sufficiently to form visible colonies. CO2 monitoring
is required in all CO2 incubators, including those that adjust gas flow to maintain a set CO2 level,
to ensure that the environment is within an acceptable range for CO2 content. It is acceptable
to monitor and record CO2 levels from digital readouts; however, the laboratory must verify
that the readout is accurate (by initial calibration, Fyrite, or other calibrated CO2 meter). The
frequency of verification of the digital readout must be defined in the laboratory's equipment
quality control procedure and should be performed, at minimum, at the frequency recommended
by the manufacturer.
MIC.21815 Campylobacter Incubation Conditions QC Phase I

Campylobacter incubation conditions are checked using QC organisms or other
appropriate methods to ensure adequate environmental conditions to support growth of
Campylobacter jejuni.
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BACTERIOLOGY SUSCEPTIBILITY TESTING

● Sampling of susceptibility test, QC and reporting policies and procedures
● Sampling of susceptibility QC records
● Susceptibility test set-up (standardized inoculum, pure culture)
● How does your laboratory work with the pharmacy and medical staff to determine
policies for reporting of antimicrobial agents?
MIC.21820 Susceptibility Testing - Pure Cultures Phase II

Only isolated colonies or pure cultures are used for performance of antimicrobial
susceptibility testing (i.e. susceptibility testing is not performed on mixed cultures).
✓ Written procedure describing the use of isolated colonies or of pure cultures for susceptibility
testing, including the use of purity plates
**REVISED** 08/17/2016
MIC.21910 Susceptibility Test QC Frequency Phase II
For antimicrobial susceptibility testing by either disk or gradient diffusion strips or broth
dilution (MIC) methods, quality control organisms are tested with each new lot number or
shipment of antimicrobials or media before or concurrent with initial use, and each day
the test is performed thereafter.
NOTE: If multiple instruments are used for automated MIC testing, QC testing should be rotated
equally among all testing instruments. The frequency of QC testing may be reduced to weekly
(including the testing of new lots or batches of antimicrobials or media) if the laboratory director
approves the use of an individualized quality control plan (IQCP), including all required elements
of IQCP, and the laboratory has records of satisfactory performance with daily QC tests as
suggested by CLSI Standards. If the laboratory has multiple instruments for automated MIC
testing, each individual instrument must be included as part of the IQCP. Please refer to the
Individualized Quality Control Plan section of the All Common Checklist for the requirements for
implementation and ongoing monitoring of an IQCP. For this purpose, satisfactory performance is
defined as follows:
1. There are records that all QC organisms were tested for 20 or 30 consecutive test
days, and
2. For each drug/microorganism combination, no more than 1 of 20 or 3 of the 30
values (zone diameter or MICs) may be outside the accepted QC ranges. These
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accepted QC ranges may be those defined in the current CLSI guidelines or

commercial device instructions or may be established by the laboratory
Or
1. There are records that all QC organisms were tested in triplicate (using separate
inoculum suspensions) for 5 consecutive test days
2. For each drug/microorganism combination, no more than 1 of the 15 values (zone
diameter or MICs) may be outside the accepted QC range
3. If 2 or 3 values are outside the accepted QC range during testing of 15 replicates,
daily QC testing must be continued and performed in triplicate (using separate
inoculum suspensions) for another 5 consecutive test days
4. For each drug/microorganism combination, no more than 4 of the 30 values (zone
diameter or MICs) may be outside the accepted QC range
When a result is outside the accepted QC range during weekly QC testing, refer to the most
recent CLSI Standards for the required corrective action.
If the laboratory performs QC on antimicrobial screening tests as defined by the CLSI Standard
and manufacturer's instructions do not require QC on each day the test is performed, the
laboratory must have an IQCP that meets all requirements defined in the All Common Checklist.
✓ Records of susceptibility QC results at defined frequency and meeting defined acceptability
criteria
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7167 [42CFR493.1261(b)(1)]
2) Clinical and Laboratory Standards Institute (CLSI). Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria; Approved
Standard—Eighth Edition. CLSI document M11-A8 (ISBN 1-56238-626-3). Clinical and Laboratory Standards Institute, 940 West
Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2012.
3) Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Disk Susceptibility Tests; Approved Standard
12th ed. CLSI Document M02-A12. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
4) Clinical and Laboratory Standards Institute. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically;
Approved Standard 10th ed. CLSI Document M07-A10. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
5) Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing; 27th ed. CLSI Document
M100-S27. Clinical and Laboratory Standards Institute, Wayne, PA; 2016.
6) Clinical and Laboratory Standards Institute. Verification of Commercial Microbial Identification and Antimicrobial Susceptibility Testing
Systems, 1st ed. CLSI Document M52-ED1. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
MIC.21930 Susceptibility Test Endpoint Determination Phase II

For antimicrobial susceptibility testing systems, there are written criteria for measuring
and determining the MIC endpoint or zone size.
NOTE: The laboratory may use CLSI criteria, but the use of other validated criteria, such as the
FDA or European Committee on Antimicrobial Susceptibility Testing (EUCAST) is acceptable.
REFERENCES
th
12 ed. CLSI Document M02-A12. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
th
Approved Standard 10 ed. CLSI Document M07-A10. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
MIC.21940 Standardized Inoculum Phase II

The inoculum used for antimicrobial susceptibility testing (i.e. inoculum size) is controlled
using a turbidity standard or other acceptable method.
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NOTE: Antibiotic susceptibility may be substantially affected by inoculum size.

✓ Written procedure for standardizing susceptibility inoculum
REFERENCES
th
12 ed. CLSI Document M02-A12. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
th
Approved Standard 10 ed. CLSI Document M07-A10. Clinical and Laboratory Standards Institute, Wayne, PA; 2015.
**REVISED** 08/21/2017
MIC.21943 Selection of Antimicrobial Agents to Report Phase II
There are written policies to ensure that only antimicrobial agents appropriate for the
organism and body site are routinely reported.
NOTE: The microbiology department should consult with the medical staff and pharmacy to
develop a list of antimicrobial agents to be reported for specific organisms isolated from various
body sites. These lists may be based on the CLSI recommendations provided in the M100 Table
1, which suggests those agents that might be reported routinely (Group A) and that might be
reported selectively (Group B). Selective reporting of antimicrobial agents should help improve
the clinical relevance of antimicrobial reporting and help minimize overuse of broad-spectrum
agents that might result in selection of multi-resistant organisms.
The antimicrobial reporting policy should include antibacterial, antifungal, and antimycobacterial
agents tested in the laboratory. Policies should be reviewed with the stakeholders involved in the
antimicrobial stewardship in the institution annually and records of the review should be available
in the laboratory. The same policies should be used in reporting proficiency testing susceptibility
results, particularly for isolates from cerebrospinal fluid and urine.
✓ Antimicrobial reporting policy AND
✓ Patient reports with reporting of antimicrobial agents for different body sites following written
policy AND
✓ Records of annual antimicrobial reporting policy review by the antimicrobial stewardship
committee AND
✓ Proficiency testing susceptibility results following written policy
REFERENCES
1) Poulter MD, Hindler JF. Challenges in Antimicrobial Susceptibility Testing and Reporting. Lab Med, November, 2002:11:33
3) CDC. Core Elements of Hospital Antibiotic Stewardship Programs. Atlanta, GA: US Department of Health and Human Services,
CDC; 2014. Available at http://cdc.gov/getsmart/healthcare/implementation/core-elements.html.
MIC.21944 Testing and Reporting Supplemental Antimicrobial Agents Phase I

There are written policies for testing supplemental agents when needed on isolates
resistant to routinely tested antimicrobial agents.
NOTE: The policy may include submission of isolates to an outside referral laboratory if testing is
not performed onsite.
✓ Patient testing reports demonstrating additional antimicrobial testing or referral
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MIC.21946 Cumulative Susceptibility Data Phase I

For hospital based microbiology laboratories, cumulative antimicrobial susceptibility test
data are maintained and reported to the medical staff at least yearly.
REFERENCES
1) Clinical and Laboratory Standards Institute. Analysis and Presentation of Cumulative Antimicrobial Susceptibility Test Data; Approved
th
Guideline. 4 ed. CLSI document M39-A4. Clinical and Laboratory Standards Institute, Wayne, PA, 2014
MIC.21950 Inconsistent Antimicrobial Results Phase I

There is a written policy to address unusual or inconsistent antimicrobial testing results.
NOTE: Acceptable results derived from testing QC strains do not guarantee accurate results
with all patient isolates. Results from testing patient isolates must be reviewed and unusual or
inconsistent results should be investigated to ensure accuracy. Expert software can identify
unusual or inconsistent results that might be due to technical errors and to identify emerging
resistance. Each laboratory should have a policy such as that provided by CLSI for confirming
unusual or inconsistent results. In some cases, it may be necessary to repeat susceptibility
testing and/or identification procedures to confirm initial results. This may involve using
alternative testing methods or sending the isolate to a referral laboratory. Some examples
include:
1. Escherichia coli resistant to carbapenems

2. Klebsiella spp. susceptible to ampicillin
3. Staphylococcus aureus resistant to vancomycin
✓ Records of investigation for unusual/inconsistent results
REFERENCES
PROCEDURES AND TESTS
● Sampling of bacteriology test procedures of different source types
● Sampling of patient worksheets/records
● Sampling of patient reports
● How does your laboratory evaluate culture findings?
● Follow a specimen from evaluation of culture findings, recording of results and

reporting. Determine if practice follows test procedures.
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ROUTINE PROCEDURES: The following requirements define minimum standards for evaluation of routine
bacterial cultures. The outlined procedure (media used and incubation conditions) permits recovery of bacteria
expected in the type of specimen used. This does not preclude the use of screening cultures (limited studies)
and should not be construed to mean that all routine cultures require special media. Special media should be
available if needed.
RESPIRATORY SPECIMENS
Routine procedures from acceptable respiratory cultures should allow the isolation of Streptococcus
pneumoniae, and Haemophilus species.
MIC.22100 Sputum Gram Stain Phase I

A gram-stained smear is performed routinely on expectorated sputa to determine
acceptability of a specimen for bacterial culture and as a guide for culture workup.
✓ Policy defining acceptable specimens
REFERENCES
1) Valenstein PN. Semiquantitation of bacteria in sputum Gram stains. J Clin Microbiol. 1988;26:1791-1794
2) Wilson ML. Clinically relevant, cost-effective clinical microbiology. Strategies to decrease unnecessary testing. Am J Clin Pathol.
1997;107:154-165
3) Church DL. Are there published standards for Gram stain results for sputa to establish specimen quality for culture? College of
American Pathologists CAP Today. 2000;14(4):97
5) Leber, AL (ed). Clinical Microbiology Procedures Handbook. 4th ed. Washington DC: ASM Press; 2016.
MIC.22110 Unacceptable Sputum Specimens Phase I

Specimens deemed unacceptable by Gram stain review are not cultured for routine
bacteria (or cultured only by special request) and the health care provider or submitting
laboratory is notified so another specimen can be collected without delay, if clinically
indicated.
NOTE: It is suggested that the laboratory notify an appropriate caregiver about an inadequate
specimen even when specimens are submitted from an outpatient setting, or submitted to
a referral laboratory. Notification can be by phone or computer report. The laboratory may
implement written agreements with particular providers or submitting laboratories defining
policies for handling sputum samples.
✓ Records of specimen rejection such as rejection log or patient report
REFERENCES
1) Bartlett RC. Medical microbiology: quality cost and clinical relevance. New York, NY: Wiley, 1974:24-31
2) Carroll KC. Laboratory Diagnosis of Lower Respiratory Tract Infection: Controversy and Conundrums. J Clin Microbiol.
2002;3115-3120
**REVISED** 08/17/2016
MIC.22140 Group A Streptococcus Direct Antigen Detection Phase I
If group A Streptococcus direct antigen testing is performed on pediatric patients,
confirmatory testing is performed on negative samples.
NOTE 1: Policies must be established for the use of cultures or other confirmatory tests
on pediatric specimens that test negative when using antigen detection methods or if the
manufacturer's guidelines include recommendations for culture follow-up. These policies should
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take into account the sensitivity of the assay in use, the age and clinical presentation of the
patient, and other factors.
NOTE 2: Direct antigen tests should be performed and reported in a timely fashion, since their
principal advantage (compared to culture) is rapid turn-around-time.
REFERENCES
1) Shulman S, Bisno A, Clegg H, et al. Clinical Practice Guideline for the Diagnosis and Management of Group A Streptococcal
Pharyngitis: 2012 Update by the Infectious Diseases Society of America. Clin Infect Dis. 2012;55(10). doi: 10.1093/cid/cis629.
URINE SPECIMENS
MIC.22200 Urine Colony Count Phase II

Quantitative cultures (colony counts) are performed.
NOTE: Urine cultures should include an estimate of CFU/volume.

✓ Written procedure for colony counts
REFERENCES
1) Susan E. Sharp, et al, Cumitech 2C: Laboratory Diagnosis of Urinary Tract Infections, 2009. ASM Press, Washington, DC
MIC.22210 Urine Culture Procedure Phase II

The media and procedures used permit the isolation and identification of both gram-
positive and gram-negative bacteria.
NOTE: This does not require the use of gram-positive selective media.
REFERENCES
1) Susan E. Sharp, et al, Cumitech 2C: Laboratory Diagnosis of Urinary Tract Infections, 2009. ASM Press, Washington, DC
GENITAL SPECIMENS
MIC.22273 Group B Streptococcus Screen Phase II

Group B streptococcus screens from pregnant women are collected and cultured in
accordance with the current guidelines.
NOTE: Universal prenatal screening for vaginal and rectal Group B streptococcal (GBS)
colonization of all pregnant women at 35-37 weeks gestation is recommended. The optimum
specimen for this test is a vaginal/rectal swab and results may be compromised if only a
vaginal swab is submitted. Detection of GBS in urine cultures in this population should also be
addressed. Procedures for collecting and processing clinical specimens for GBS culture and
performing susceptibility testing to clindamycin and erythromycin for highly penicillin allergic
women are also included in the guidelines.
REFERENCES
1) Center for Disease Control and Prevention, 2010. Prevention of perinatal Group B streptococcal disease MMWR 59(RR-10);1-32
MIC.22280 Bacterial Vaginosis Phase I

When Gram stains are performed to make the laboratory diagnosis of bacterial vaginosis,
the smear is scored and interpreted according to published criteria.
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NOTE: Culture should not be used for the diagnosis of bacterial vaginosis. Bacterial vaginosis
(BV) is a syndrome involving a shift in the concentrations of aerobic and anaerobic flora of the
genitourinary tract flora from a predominant presence of Lactobacillus sp. to that of a mixture
of anaerobes, Gardnerella vaginalis and other gram-negative bacteria. Culturing for a particular
organism, such as Gardnerella vaginalis, or any single organism or combination of organisms is
not specific for the diagnosis of BV. Use of a scored Gram stain that demonstrates whether there
has been a shift in the vaginal flora from predominantly gram-positive Lactobacillus to a gram-
negative flora has been shown to correlate well with the Amsel criteria for the diagnosis of BV.
The primary reason for performing a Gram stain on vaginal secretions is to diagnose bacterial
vaginosis.
REFERENCES
1) Nugent RP, Krohn MA, Hillier SL. 1991. JCM 29;297-301
2) Forsum U, Hallen A, Larsson PG. Bacterial Vaginosis - a laboratory and clinical diagnostic enigma. Review article III. APMIS 113:
153-61. 2005
MIC.22285 Genital Pathogens Phase II

Written procedures are established to ensure the detection of genital pathogens such as
Neisseria gonorrhoeae.
STOOL SPECIMENS
MIC.22330 Clostridium difficile Phase II

There are written policies for the testing of stool for the detection and reporting of C.
difficile or its toxins.
REFERENCES
1) Novak-Weekley SM, et al. Clostridium difficile testing in the Clinical Laboratory by Use of Multiple Testing Algorithms. Journal of
Clinical Microbiology 2010; 48:889-893
2) Eastwood K, et al. Comparison of Nine Commercially Available Colostridium difficile Toxin Detection Assays, a Real-Time PCR
Assay for C. difficile tcdB and a Glutamate Dehydrogenase Detection Assay to Cytotoxin Testing and Cytotoxigenic Culture Methods.
Journal of Clinical Microbiology 2009; 47:3211-3217
3) Peterson LR and Robicsek A. Does my Patient have Clostridium difficile Infection? Annals of Internal Medicine 2009; 151:176-178
MIC.22336 Stool Specimen Reporting Phase I

The final report for stool cultures submitted for routine bacterial pathogen examination
lists the organisms for which the specimen was cultured (e.g. Salmonella, Shigella,
Vibrio).
NOTE:
1. It is inappropriate to report “No enteric pathogens isolated.” The report should list the
organisms whose presence was specifically sought (e.g. No Salmonella, Shigella, or
Campylobacter isolated).
2. When indicated, tests to detect Shiga toxin-producing E. coli (STEC) should be
available at a referral laboratory if not performed onsite.
REFERENCES
1) Gilligan PH, et al. Laboratory diagnosis of bacterial diarrhea. Cumitech 12A, 1992. ASM Press; Washington DC
MIC.22410 Stool Culture Enrichment/Selective Media Phase I

Appropriate methods are used routinely to recover enteric pathogens.
NOTE: Enrichment media may be used in addition to selective plating media to enhance
recovery of pathogens, which may be present at low numbers.
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MIC.22440 Stool Specimen Number/Timing Phase I

There are written policies for the number and/or timing of collection of stool specimens
submitted for routine bacterial testing.
NOTE: The laboratory should consider developing policies with its clinicians for the number and/
or timing of collection of stool specimens submitted for routine bacterial testing. Suggestions
made by the authors of a 1996 CAP Q-Probes study (Valenstein et al) include:
1. Accept no more than two specimens/patient without prior consultation with an

individual who can explain the limited yield provided by additional specimens
2. Do not accept specimens from inpatients after the third hospital day, without prior
consultation
3. Test stool for Clostridium difficile toxin for all patients with clinically significant
diarrhea and a history of antibiotic exposure. Consider C. difficile testing as an
alternative to routine microbiologic studies for inpatients who have test requests for
routine enteric pathogens
4. Positive test results for Clostridium difficile do not correlate well with disease in
young children. Follow manufacturer's guidelines for guidance on the testing of
pediatric patients.
These recommendations are for diagnostic testing. Different policies may apply to tests ordered
for follow-up.
REFERENCES
1) Yannelli B, et al. Yield of stool cultures, ova and parasite tests, and Clostridium difficile determinations in nosocomial diarrhea. Am J
Infect Control. 1988;16:246-249
2) Siegel DL, et al. Inappropriate testing for diarrheal diseases in the hospital. JAMA. 1990;263:979-982
3) Asnis DS, et al. Cost-effective approach to evaluation of diarrheal illness in hospitals. J Clin Microbiol. 1993;31:1675
4) Fan K, et al. Application of rejection criteria for stool cultures for bacterial enteric pathogens. J Clin Microbiol. 1993;31:2233-2235
5) Valenstein P, et al. The use and abuse of routine stool microbiology. A College of American Pathologists Q-probes study of 601
institutions. Arch Pathol Lab Med. 1996;120:206-211
1997;107:154-165
7) Blackman E, et al. Cryptosporidiosis in HIV-infected patients: diagnostic sensitivity of stool examination, based on number of
specimens submitted. Am J Gastroenterol. 1997;92:451-453
8) Wood M. When stool cultures from adult inpatients are not appropriate. Lancet. 2001;357:901-902
9) Bauer, TM, et al. Derivation and validation of guidelines for stool cultures for enteropathogenic bacteria other than Clostridium difficile
in hospitalized patients. JAMA. 2001;285:313-319
CEREBROSPINAL & OTHER BODY FLUID SPECIMENS
MIC.22495 Centrifugation of Body Fluids Phase I

If only plated media are used for sterile body fluids, fluid is centrifuged and the sediment
used to inoculate media unless the entire specimen is plated.
NOTE: When inadequate volume is received, the report should note that the culture results
may be compromised by the limited volume of specimen received. Equivalent methods are
acceptable, if validated by the laboratory.
REFERENCES
1) Versalovic, James. Manual of Clinical Microbiology - 10th Edition. Washington, DC: ASM Press. 2011
MIC.22500 CSF Processing Phase II

CSF samples for culture are processed immediately on receipt.
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NOTE: Bacterial meningitis is a critical condition that requires immediate attention. Samples must
be processed upon receipt when meningitis is suspected. The laboratory may choose to handle
surveillance cultures, e.g. involving neurosurgical implants, differently.
✓ Policy and procedure for CSF processing AND
✓ Culture log or patient records
MIC.22520 CSF Media/Incubation Phase II

The procedure (media and incubation conditions) permits recovery of fastidious bacteria
expected in this type of specimen (N. meningitidis, S. pneumoniae, H. influenzae).
MIC.22550 CSF Back-Up Cultures Phase II

If bacterial antigen-detection methods are used, back-up cultures are performed on both
positive and negative CSF specimens.
NOTE: Total dependence on a bacterial antigen test for the diagnosis of bacterial meningitis
does NOT meet accreditation requirements. Meningitis may be caused by bacteria not detected
by the antigen tests. Thus, culture is essential for proper evaluation of bacterial meningitis,
and must be performed on the patient specimen - if not performed onsite by the laboratory, the
inspector must seek evidence that a culture has been performed in a referral laboratory.
✓ Written policy stating that CSF cultures are performed in conjunction with bacterial antigen
tests OR policy describing testing at another location AND
✓ Records of back-up CSF cultures performed on-site OR records indicating that cultures
are performed at another location OR records that the order for CSF bacterial antigen was
blocked by the computer due to no order for a culture
REFERENCES
1) Forward KR. Prospective evaluation of bacterial antigen detection in cerebral spinal fluid in the diagnosis of bacterial meningitis in a
predominantly adult hospital. Diagn Micro Infect Dis. 1988;11:61-63
2) Maxson S, et al. Clinical usefulness of cerebrospinal fluid bacterial antigen studies. J Pediat. 1994; 125:235-238
3) Finlay FO, et al. Latex agglutination testing in bacterial meningitis. Arch Dis Child. 1995;73:160-161
4) Rathore MH, et al. Latex particle agglutination tests on the cerebrospinal fluid. A reappraisal. J Florida Med Assoc. 1995;82:21-23
5) Kiska DL, et al. Quality assurance study of bacterial antigen testing of cerebrospinal fluid. J Clin Micro. 1995;33:1141-1144
6) Perkins MD, et al. Rapid bacterial antigen detection is not clinically useful. J Clin Micro. 1995;33:1486-1491
BLOOD CULTURES
MIC.22600 Blood Culture System Phase II

The blood culture system in use is designed to recover both aerobic and, when indicated
or if intended to be part of the routine procedure, anaerobic organisms.
NOTE: This criterion is not intended to imply that anaerobic cultures must be performed on
all blood cultures if circumstances where anaerobic cultures are not indicated are specifically
delineated (e.g. on neonates where volume is of concern).
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Principles and Procedures for Blood Cultures; Approved Guideline. CLSI
Document M47-A (ISBN 1-56238-641-7). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne, PA
19087-1898 USA, 2007.
2) Baron EJ, et al. Blood Cultures IV, Cumitech 1C, 2005, ASM Press; Washington, DC
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**REVISED** 08/17/2016
MIC.22610 Manual Blood Culture Systems Phase II
For non-automated systems, macroscopically negative aerobic blood cultures are stained
and/or subcultured within 12-48 hours of incubation.
✓ Records of staining and/or subculture of macroscopically negative cultures
REFERENCES
19087-1898 USA, 2007.
MIC.22620 Blood Culture Examination Phase II

Blood cultures are examined (macroscopically if manual method) for evidence of growth
at least twice daily for the first two days of incubation, then at least daily for the remainder
of the incubation period.
NOTE: The time to detection of positive blood cultures, whether processed by manual or
automated methods, depends on the schedule of inspection for evidence of growth. The means
of the inspection may include visual examination, gram staining, subculturing, or electronic
analysis by continuous monitoring instruments. Because most significant positive blood cultures
may be detected within 48 hours of incubation, it is recommended that blood cultures be
examined for evidence of growth at least two times on the first two days of incubation, then at
least once daily through the remainder of the laboratory's routine incubation period.
✓ Patient records/worksheet with result of examination for manual methods at defined
frequency
REFERENCES
19087-1898 USA, 2007.
MIC.22630 Blood Culture Collection Phase II

Sterile techniques for drawing and handling of blood cultures are defined, made available
to individuals responsible for specimen collection and practiced.
NOTE: It is recommended that blood culture statistics, including number of contaminated

cultures, be maintained and reviewed regularly by the laboratory director. The laboratory should
establish a threshold for an acceptable rate of contamination. Tracking the contamination rate
and providing feedback to phlebotomists or other persons drawing cultures has been shown to
reduce contamination rates. Other measures to monitor include types of skin disinfection, volume
of blood drawn, number of culture sets drawn, number of single cultures and line draws.
REFERENCES
1) Gibb AP, et al. Reduction in blood culture contamination rate by feedback to phlebotomists. Arch Pathol Lab Med. 1997;121:50-507
2) Schifman RB, et al. Blood culture contamination. A College of American Pathologists Q-probes study involving 640 institutions and
497 134 specimens from adult patients. Arch Pathol Lab Med. 1998;122:216-221
3) DesJardin JA, et al. Clinical utility of blood cultures drawn from indwelling central venous catheters in hospitalized patients with
cancer. Ann Int Med. 1999;131:641-647
4) Ernst DJ. Controlling blood culture contamination rates. Med Lab Observ. 2000;32(5):36-47
5) Ruge DG, et al. Reduction of blood culture contamination rates by establishment of policy for central intravenous catheters. Lab Med.
2002;33:797-800
19087-1898 USA, 2007.
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**REVISED** 08/17/2016
MIC.22640 Blood Culture Volume Phase I
The laboratory has a written policy and procedure for monitoring blood cultures from
adults for adequate volume and providing feedback on the results to blood collectors.
NOTE: Larger volumes of blood increase the yield of true positive cultures. In adults, optimally
20 mL of blood per culture set (two bottles) should be collected for culture. The laboratory should
periodically monitor collected blood volumes and provide feedback to clinical staff. Automated
blood culture systems approved or cleared by the FDA may use smaller volumes per culture set
and are acceptable.
✓ Records of monitoring of volume at a defined frequency AND
✓ Records of feedback to the clinical staff
REFERENCES
1) Kellogg JA, et al. Justification and implementation of a policy requiring two blood cultures when one is ordered. Lab Med.
1994;25:323-330
2) Li J, et al. Effects of volume and periodicity on blood cultures. J Clin Microbiol. 1994;32:2829-2831
3) Schiffman RB, et al. Blood culture quality improvement. A College of American Pathologists Q-probes study involving 909 institutions
and 289 572 blood culture sets. Arch Pathol Lab Med. 1996;120:999-1002
4) Arendup M, et al. Diagnosing bacteremia at a Danish hospital using one early large blood volume for culture. Scand J Infect Dis.
1996;28:609-614
1997;107:154-165
6) Novis DA, et al. Solitary blood cultures. A College of American Pathologists Q-Probes study of 132778 blood culture sets in 333 small
hospitals. Arch Pathol Lab Med. 2001;125:1290-1294
19087-1898 USA, 2007.
ANAEROBIC CULTURES
**NEW** 08/17/2016
MIC.22675 Anaerobic Cultures Phase I
The laboratory has written policies defining when to culture for anaerobes.
NOTE: The policy must define criteria to determine if the submitted material is of sufficient quality
to provide an interpretable result.
If the laboratory does not perform anaerobic cultures on-site, the laboratory must refer specimens
to a referral laboratory for anaerobic culture when indicated.
**REVISED** 08/17/2016
MIC.22700 Anaerobic Cultures Phase II
There are written procedures describing how to culture anaerobic organisms when
indicated.
NOTE: For example, the minimum standards for the evaluation of deep wound cultures
require adequate procedures for the collection, recovery and identification of clinically relevant
pathogens, which includes aerobic, facultatively anaerobic, and strictly anaerobic organisms.
Anaerobic organisms may be significant isolates from other specimen types as well. Suggested
media for anaerobes include an anaerobic blood agar plate, a medium that inhibits gram-positive
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and facultative gram-negative bacilli such as KV blood agar, a differential or selective medium
such as BBE (Bacteroides bile-esculin), and a gram-positive selective medium (colistin-nalidixic
acid blood agar or phenylethyl alcohol blood agar). Provisions for adequate anaerobic incubation,
with monitoring of the anaerobic environment, must be available. If specimens are referred
to another laboratory, they must be transported in an expeditious fashion under appropriate
conditions.
REFERENCES
WOUND SPECIMENS
GAS CHROMATOGRAPHY (GC) FOR MICROBIAL IDENTIFICATION
● Sampling of gas chromatography policies and procedures
● Sampling of QC records
● Sampling of reagents (grade)

● Chromatograph pattern controls
MIC.22790 GC Calibrators/Standards Phase II

Appropriate calibrators or standards are run with each analytic batch.
NOTE: For GC, a calibrator mixture must be run approximately every tenth analysis. The
calibrator mixture must contain acids that cover the entire analysis spectrum, that is, C-10
through C-20, as well as any labile acids such as hydroxyl fatty acids.
✓ Written procedure defining frequency and content of calibrator mixtures AND
✓ Records of calibration/calibration verification with each batch
MIC.22810 GC Controls Phase II

Appropriate controls are extracted and run through the entire procedure.
NOTE: For any GC system, positive controls must be run daily and include two organisms
containing representative cellular fatty acids of all classes, i.e. saturated, unsaturated, iso,
cyclopropane and hydroxyl acids. If one is using the MIDI (or related system), the similarity
index must be >0.6 for aerobic bacteria and >0.3 for anaerobes. This represents a procedure
control, as opposed to a calibrator mixture, which is an instrument control. For all GC assays
that are used for identification of microbes, a reagent blank must be run daily to evaluate reagent
contamination and carry-over.
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REFERENCES
1) Department of Health and Human Services, Centers for Medicare and Medicaid Services. Medicare, Medicaid and CLIA programs;
CLIA fee collection; correction and final rule. Fed Register. 2003(Jan 24):5232 [42CFR493.1256]
2) Winn W et al. The anaerobic bacteria in Koneman's color atlas and textbook of diagnostic microbiology. 6th ed. Philadelphia, PA:
Lippincott Williams & Wilkins, 2006:906-912
MIC.22820 Chromatogram Controls Phase I

External chromatogram pattern controls are available.
NOTE: Patterns for known strains should be established. In addition, laboratories should have
access to the standard method manuals containing comparable chromatographic patterns for
comparison.
**REVISED** 08/17/2016
MIC.22830 GC Growth Conditions Phase II
Written policies define growth media and conditions acceptable for gas chromatography
(GC) analysis.
NOTE: Final results can be influenced by conditions of growth. For reliable results, standard
conditions of analysis must be met, including growth media.
REFERENCES
MIC.22840 Chromatographic Method Validation Phase II

The chromatographic method has been validated using known strains of bacteria,
including strains expected to be encountered in routine clinical use.
✓ Record of method validation with appropriate strains
REFERENCES
MIC.22850 Chromatographic Result Review Phase II

There is a written policy for review of chromatographic results in conjunction with other
laboratory data, prior to reporting results.
NOTE: Chromatography is only one tool for microbial identification. When results of analysis
conflict with growth characteristics, pigmentation, or the results of biochemical or molecular
testing, identification must be based on all the information available.
REFERENCES
2) Welch DF. Application of cellular fatty acid analysis. Clin Micro Rev. 1991;4:422-38
MIC.22860 Chromatographic Analysis - Pure Isolates Phase II

There are procedures to check the purity of cultures used as a source for
chromatographic analysis.
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NOTE: Results of chromatographic analysis may be unreliable if mixed cultures are tested.
If chromatography is performed on an isolate from liquid culture and an interpretable
chromatogram is obtained, it is not necessary to await the results of the purity check before
reporting results, but a purity check must still be performed.
MIC.22870 GC Reagent Grade Phase I

Reagents are of appropriate grade.
✓ Reagent logs

There are written procedures for operation and calibration of the gas chromatography
instrument.
NOTE: Basic principles of chromatographic analysis require continual monitoring of analysis

conditions, including system calibration. System problems and corrective actions must be
appropriately documented.
LABORATORY SAFETY
● Hazardous waste disposal policy
● How does your laboratory dispose of specimens and contaminated media?
NOTE TO THE INSPECTOR: The inspector should review relevant requirements from the Safety section of the
Laboratory General checklist, to assure that the bacteriology laboratory is in compliance. Please elaborate upon
the location and the details of each deficiency in the Inspector's Summation Report.
The following requirement pertains specifically to the bacteriology laboratory.
MIC.23200 Hazardous Waste Disposal Phase II

Microbiology specimen residuals and contaminated media are disposed of in a manner to
minimize hazards to all personnel handling the material.
NOTE: Sterilization or decontamination within the microbiology section before disposal is

preferred. If such material is transported before treatment, it must be placed into a leak-resistant
rigid container, and appropriately labeled.
✓ Written policy for the handling and disposal of microbiology waste
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REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Clinical Laboratory Waste Management; Approved Guideline—Third Edition. CLSI
document GP05-A3 (ISBN 1-56238-744-8). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne,
Pennsylvania 19087-1898, USA 2011.
MYCOBACTERIOLOGY
QUALITY CONTROL
● Mycobacteriology specimen collection, transport and handling policy
● Mycobacteriology reporting policy
● Sampling of patient test reports
● Sampling of mycobacteriology media/stain/reagent QC policies and procedures
● Sampling of mycobacteriology QC records
● Sampling of media (expiration date, condition, contamination)
● What is your policy for performing AFB stains on week-ends and holidays?
SPECIMEN HANDLING
MIC.31100 Specimen Collection/Transport Phase I

Specimens for mycobacterial testing are collected appropriately and transported to the
laboratory without delay.
NOTE: The laboratory should recommend collecting three sputum specimens for acid-fast
smears and culture in patients with clinical and chest x-ray findings compatible with tuberculosis.
These three samples should be collected at 8-24 hour intervals and should include at least one
first morning specimen. Specimens must be delivered to the laboratory promptly; specimens
that cannot be processed within one hour of the time of collection should be refrigerated during
transport to and storage in the laboratory prior to processing. This will decrease overgrowth with
contaminating organisms likely to be present.
Laboratories are encouraged to process acid-fast specimens in their laboratory or obtain results
from referral laboratories as soon as possible so that smear results can be available within 24
hours of collection (see MIC.31200 below).
✓ Written policy describing specimen collection and handling requirements
REFERENCES
1) Toman K. How many bacilli are present in a sputum specimen found positive by smear microscopy [Chapter 4]. In: Frieden T, ed.
Toman's tuberculosis case detection, treatment, and monitoring: questions and answers. 2nd ed. Geneva, Switzerland: World Health
Organization; 2004:11-3
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2) Siddiqui AH, Perl TM, Conlon M, Donegan N, Roghmann MC. Preventing nosocomial transmission of pulmonary tuberculosis: when
may isolation be discontinued for patients with suspected tuberculosis? Infect Control Hosp Epidemiol 2002;23:141-4
3) CDC. "Treatment of Tuberculosis: American Thoracic Society, CDC, and Infectious Diseases Society of America" MMWR 2003:52
(No.RR-11)
4) Hopewell PC, Pai M, Maher D, et al. International Standards for Tuberculosis Care. Lancet 2006;6:710-25
5) Centers for Disease Control and Prevention. Guidelines for Preventing the Transmission of Mycobacterium Tuberculosis in Health-
Care Settings. MMWR, 2005; 54(RR17);1-141.
REPORTING OF RESULTS
MIC.31200 Acid Fast Stain Results Phase I

When clinically indicated, results of acid-fast stains are reported within 24 hours of
specimen receipt by the testing laboratory.
✓ Written policy defining turnaround time for reporting acid-fast stain results
REFERENCES
1) Huebner RE, et al. Current practices in mycobacteriology: results of a survey of state public health laboratories. J Clin Microbiol.
1993;31:771-775
2) Tenover FC, et al. The resurgence of tuberculosis: is your laboratory ready? J Clin Microbiol. 1993;31:767-770
3) Woods GL, et al. Mycobacterial testing in clinical laboratories that participate in the College of American Pathologists
mycobacteriology surveys. Changes in practices based on responses to 1992, 1993, and 1995 questionnaires. Arch Pathol Lab Med.
1996;120:429-435
MIC.31220 Susceptibility Test Results Phase I

Susceptibility test results for M. tuberculosis are available in a timely manner.
NOTE: The rapid recognition of drug-resistant organisms is essential to the control of multidrug-
resistant tuberculosis. For isolates of M. tuberculosis complex, the CDC and Prevention
Laboratory work group recommends that laboratories use methods that may allow susceptibility
test results to be available within 28 days of specimen receipt. From a CAP accreditation
perspective, 28 days is a goal, not a requirement.
REFERENCES
1) Tenover FC, et al. The resurgence of tuberculosis: is your laboratory ready? J Clin Microbiol. 1993;31:767-770
2) College of American Pathologists position statement regarding rapid detection of Mycobacterium tuberculosis. Arch Pathol Lab Med.
1993;117:873
3) Woods GL, Witebsky FG. Current status of mycobacterial testing in clinical laboratories. Arch Pathol Lab Med. 1993;117:876-884
4) Huebner RE, et al. Current practices in mycobacteriology: results of a survey of state public health laboratories. J Clin Microbiol.
1993;31:771-775
5) Clinical and Laboratory Standards Institute (CLSI). Susceptibility Testing for Mycobacteria, Nocardia, and Other Aerobic
Actinomycetes; Approved Standard - Second Edition. CLSI document M24-A2. Clinical and Laboratory Standards Institute, 940
West Valley Road, Suite 1400 Wayne, PA 19087-1898, USA, 2011.
6) Woods GL, et al. Mycobacterial testing in clinical laboratories that participate in the College of American Pathologists
mycobacteriology surveys. Changes in practices based on responses to 1992, 1993, and 1995 questionnaires. Arch Pathol Lab Med.
1996;120:429-435
7) Woods GL, Witebsky FG. Susceptibility testing of Mycobacterium avium complex in clinical laboratories. Results of a questionnaire
and proficiency test performance by participants in the College of American Pathologists mycobacteriology E survey. Arch Pathol Lab
Med. 1996;120:436-439
8) Howanitz JH, Howanitz PJ. Timeliness as a quality attribute and strategy. Am J Clin Pathol. 2001;116:311-315
MEDIA

1. Sterility
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NOTE: The laboratory must have records showing that all media are sterile, able to support
growth, and are appropriately reactive biochemically. This will ordinarily require that the
use. End user quality control must be performed on media not listed on Table 2 of M22-A3 (e.g.
dermatophyte test medium), regardless of the exempt status.
This checklist requirement does not apply to commercially prepared additives that are
reconstituted when added to mycobacterial media.
Laboratories receiving media from media suppliers must have a copy of the CLSI/NCCLS
Standard M22-A3 as a reference source. The media supplier must provide records showing
that the quality control activities meet the CLSI/NCCLS Standard M22-A3, or are otherwise
equivalent. The laboratory director may wish to have a signed contractual arrangement with his/
her selected media supplier to cover all expected quality control and documentation thereof.
✓ Written procedure for QC on new lot numbers or shipments of purchased medium AND
applicable AND
REFERENCES

appropriate sample of each medium and additive is checked before or concurrent with
initial use for each of the following:
1. Sterility
previous batches
✓ Records of media QC for laboratory-prepared media and additives
REFERENCES
1) Sharp SE, et al. Lowenstein-Jensen media. No longer necessary for mycobacterial isolation. Am J Clin Pathol. 2000:113:770-773
MIC.31460 Media Visual Examination Phase II

All media are in satisfactory condition (adequately hydrated, tubed media not dried or
loose from sides).
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CONTROLS AND STANDARDS
MIC.31640 AFB Stain QC Phase II

Acid-fast bacillus stains are checked each day of use with appropriate positive and
negative controls, and results recorded.
REFERENCES

Fluorescent stains are checked with positive and negative controls each time of use and
results recorded.
REFERENCES
MIC.31670 Nucleic Acid Probe QC Phase II

If nucleic acid probes are used for identification of mycobacteria grown in culture,
appropriate positive and negative controls are tested on each day of use.
✓ Records of nucleic acid probe QC at defined frequency
REFERENCES
1) Woods GL. Molecular methods in the detection and identification of mycobacterial infections. Arch Pathol Lab Med.
1999;123:1002-1006
MIC.31680 Susceptibility QC Phase II

If the laboratory performs susceptibility testing of M. tuberculosis, a control strain
susceptible to all antimycobacterial agents is run each week of patient testing, and with
each new batch/lot number of media and antimicrobial agents.
✓ Records of routine and new lot QC results at defined frequency
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3708 [42CFR493.1262(b)]
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● Sampling of mycobacteriology test procedures/identification schemes
● Sampling of mycobacteriology QC policies and procedures
● Sampling of mycobacteriology QC records
● Sampling of HPLC policies and procedures
● Sampling of HPLC QC records
● New HPLC column verification
● Chromatograph pattern controls

● Sampling of reagents (storage, grade, solvent purity)
● What specimens are concentrated before AFB examination and culture?
● Follow a positive patient worksheet from receipt and processing to identification and
reporting
RAPID METHODS
The College of American Pathologists encourages laboratories in areas of the country where the incidence
of tuberculosis has increased over the past several years and laboratories in other parts of the country that
have experienced an increased rate of recovery of mycobacteria to utilize the most rapid and reliable methods
available for detection and identification of mycobacteria, especially M. tuberculosis, and the most rapid and
reliable methods available for susceptibility testing of isolates of M. tuberculosis.
**REVISED** 08/17/2016
MIC.32100 Fluorochrome Stain Phase II
Fluorochrome staining is performed on mycobacterial smears prepared from primary
respiratory specimens, either in the laboratory or by the referral laboratory.
NOTE: Such smears are easier to read than those stained with a conventional carbol-fuchsin
based stain. Fluorescing organisms stand out prominently against the background of the smear,
and the smears can be examined at a lower power than conventionally-stained smears, so that
a larger amount of material can be examined in a given period of time. As with the interpretation
of Ziehl-Neelsen- and Kinyoun-stained smears, expertise is needed for interpretation of smears
stained with a fluorescent stain; not everything that fluoresces in such a stain is necessarily a
mycobacterium. Particularly when only a few organism-like structures are seen, it is important to
pay careful attention to their morphology before interpreting them as mycobacteria.
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This requirement does not apply to laboratories outside of the United States where local
regulations prevent fluorochrome staining.
✓ Written policy for including fluorochrome staining on primary respiratory specimens for
mycobacterial culture OR written policy for referral of specimens to a referral laboratory for
fluorochrome staining AND
✓ Patient reports/worksheets with fluorochrome stain results OR referral laboratory reports with
results
REFERENCES
1) Narain R, et al. Microscopy positive and microscopy negative cases of pulmonary tuberculosis. Am Rev Resp Dis. 1971;103:761-773
2) Lipsky BA, et al. Factors affecting the clinical value of microscopy for acid-fast bacilli. Rev Infect Dis. 1984;6:214-222
3) Witebsky FG. Q&A. College of American Pathologists CAP TODAY, 1999;13(1):72
4) Somoskovi A, et al. Lessons from a proficiency testing event for acid-fast microscopy. Chest. 2001;120:250-257
MIC.32140 Rapid Method Phase I

Nucleic acid probes, chromatography, the NAP test, or other rapid method (e.g. nucleic
acid amplification or sequencing) is employed for identification of mycobacterial isolates.
✓ Written policy defining method(s) in use for identification of mycobacterial isolates
REFERENCES
1) Smith MB, et al. Evaluation of the enhanced amplified Mycobacterium tuberculosis direct test for direct detection of Mycobacterium
tuberculosis complex in respiratory specimens. Arch Pathol Lab Med. 1999;123:1101-1103
2) Driscoll JR, et al. How and why we fingerprint tuberculosis. RT J Resp Care Pract. 2001:Feb/Mar
CONCENTRATION, INOCULATION, INCUBATION
MIC.32200 AFB Concentration Phase II

There is a written policy defining those specimens (e.g. sputum) requiring concentration
before AFB smear examination and culture.
MIC.32250 Specimen Inoculation Phase I

Specimens (other than blood) are routinely inoculated on media that support optimal
growth of the majority of clinically relevant mycobacterial species.
NOTE: The use of two types of media (for specimens other than blood), including one liquid
medium (when possible) or a comparable culture method, is recommended for optimal isolation
of mycobacteria.
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Laboratory Detection and Identification of Mycobacteria; Approved Guideline.
CLSI document M48-A (ISBN 1-56238-659-7). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400,
Wayne, Pennsylvania 19087-1898 USA, 2008.
CULTURES
Laboratories providing complete identification must provide a sufficient variety of differential tests to accurately
identify and differentiate the different types of mycobacteria, including temperature growth requirements and
photoreactivity studies. Laboratories not providing complete identification are encouraged to at least provide
photoreactivity studies.
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MIC.32320 Incubation Temperature Phase II

Mycobacterial cultures are maintained at 35 to 37°C.
NOTE: The optimal incubation temperature for most mycobacterial specimens is 35 to 37°C.
Exceptions to this include specimens obtained from skin or soft tissue suspected to contain M.
marinum (incubate at 30 to 32°C) or M. xenopi (incubate at 42°C). These specimens should be
held at 35 to 37°C in addition to the lower or higher temperature.
✓ Temperature records
DIFFERENTIAL BIOCHEMICAL PROCEDURES
MIC.32420 Differential Biochemical Test Phase II

Differential biochemical tests are appropriate for the extent and manner of mycobacterial
identification.
NOTE: The number and types of biochemical tests needed depend upon (a) the extent to
which mycobacteria are identified (e.g. "Mycobacterium kansasii" or "photochromogen"), (b)
the particular species which a laboratory attempts to identify (e.g. does it attempt to identify
Mycobacterium terrae complex, or the species and subspecies of the Mycobacterium chelonae-
Mycobacterium fortuitum complex), and (c) the degree to which biochemical testing is ancillary to
other methods such as nucleic acid probes and HPLC. Useful biochemical tests include, but are
not limited to, arylsulfatase, 68° C catalase, semiquantitative catalase, iron uptake, MacConkey
agar, 5% NaCl, niacin accumulation, nitrate reductase, Tween 80 hydrolysis, and urease. These
tests are particularly useful for the following identifications and discriminations:
TEST UTILITY
Arylsulfatase Helps distinguish pathogenic from non-pathogenic rapid growers;
also useful for M. marinum, M. szulgai, M. xenopi, M. triviale.
68°C catalase Helpful for identification of M. tuberculosis
Semiquantitative Helpful in certain circumstances. M.
catalase tuberculosis complex, MAC, M. xenopi, and a few other species
produce <45 mm of bubbles.
Iron uptake Helps distinguish M. chelonae from M. fortuitum.
MacConkey agar Helps with identification of rapid growers.
5% NaCl Helps with identification of rapid growers and M. triviale.
Niacin accumulation Helps with identification of M. tuberculosis, M. simiae, some strains
of M. bovis.
Nitrate reductase Helpful in identifying many mycobacterial species.
Tween 80 hydrolysis Helps distinguish some usually pathogenic from some usually non-
pathogenic mycobacterial species.
Urease Helpful in identifying many mycobacterial species.
✓ Written procedure detailing tests performed and identification scheme appropriate for the
extent of testing
REFERENCES
1) Kent PT, Kubica GP. Public health mycobacteriology: a guide for the level III laboratory. Atlanta, GA: US Department of Health and
Human Services, Centers for Disease Control, 1985
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MIC.32480 Biochemical Test QC Phase II

All biochemical tests employed are checked each day of use with appropriate positive and
negative controls and results recorded.
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3708 [42CFR493.1262(a)]
HIGH PERFORMANCE LIQUID CHROMATOGRAPHY

(HPLC) FOR MICROBIAL IDENTIFICATION
MIC.32518 HPLC Calibrators/Standards Phase II

Appropriate calibrators or standards are run with each analytic batch.
NOTE: Either calibration standards or organisms of known identity must be run with each analytic
batch, and criteria must exist for acceptance of runs based on mobility of internal standards,
ability to identify significant peaks, baseline noise, peak symmetry of internal standards, detection
of low-quantity peaks, and similar criteria.
✓ Written policy defining calibrators/standards appropriate for the test system used AND
✓ Records of calibration/calibration verification with each batch
REFERENCES
1) Department of Health and Human Services, Centers for Disease Control and Prevention, Steering Committee of the HPLC User's
Group. Standardized Method for HPLC Identification of Mycobacteria, 1996
MIC.32556 HPLC Controls Phase II

Appropriate controls are extracted and run through the entire procedure.
NOTE: Control organisms must be extracted and carried through the entire procedure with each
run or batch. Appropriate positive (e.g. mycobacterial) and negative controls (organisms such as
Candida from which no mycolic acids are expected) must be included with each run.
REFERENCES
MIC.32594 Chromatogram Controls Phase I

External chromatogram pattern controls are available.
NOTE: Patterns for known strains should be established in those laboratories using HPLC. In
addition laboratories should have access to the standard method manuals containing comparable
chromatographic patterns for comparison.
REFERENCES
MIC.32632 Column Verification Phase II

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New columns are verified for performance before use.
NOTE: Column verification must include assessment of flow, consistency, and carryover. If
the HPLC-method interpretive software uses a peak-naming table, it must be calibrated with
each change of column. Generally the basic performance of new columns is certified by the
manufacturer. HPLC analysis requires columns be equilibrated with about 10 column volumes of
solvent followed with a blank run to test pressure and solvent flow.
✓ Written procedure for column verification AND
✓ Records of column verification
REFERENCES
MIC.32670 Column/Detector Monitoring Phase II

The performance of the column and detector are monitored on each day of use.
NOTE: Unextracted standard organisms and extracted calibrators or controls, typically containing
a range of mycolic acids (or other appropriate targets) of known relative retention times, may
be analyzed to monitor critical aspects of HPLC performance. Appropriate criteria for evaluating
such parameters as retention time of specific standards, relative retention compounds time,
separation of closely eluting peaks of interest, detection of known low-quantity peaks, column
pressure, chromatography quality and detector response should be established and monitored.
Column temperatures and pump pressures are monitored with each run to ensure they met
specified criteria for analysis. The column and detector operations are monitored with a blank run
prior to use and during batch runs. Positive and negative control samples supplement the blank
run when samples are analyzed.
✓ Records for column and detector monitoring at defined frequency
REFERENCES
MIC.32708 Carryover Detection Phase II

There is a written procedure for the detection and evaluation of potential carryover.
NOTE: No matter what type of injection is used, the written procedure must address criteria for
the evaluation of potential carryover from a preceding elevated (high concentration) sample to
the following sample, either periodically, or in each analytical batch analysis.
✓ Records of reassessment of samples with potential carryover
REFERENCES
1) Clinical and Laboratory Standards Institute. Preliminary Evaluation of Quantitative Clinical Laboratory Methods; Approved Guideline.
rd
3 ed. CLSI Document EP10-A3-AMD. Clinical and Laboratory Standards Institute, Wayne, PA; 2014.
2) Society of Forensic Toxicologists/American Academy of Forensic Sciences. Forensic Toxicology Laboratory Guidelines. 2002;
8.2.8:13
MIC.32746 HPLC Growth Media Phase II

The laboratory policies define which growth media may be used for organisms to be
analyzed by HPLC.
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NOTE: Final results can be influenced by conditions of growth. For reliable results, standard
conditions of analysis must be met, including growth media.
REFERENCES
MIC.32784 Peak Verification Phase II

There is a procedure for verifying calibration of the peak-naming table, if used.
NOTE: In order to insure that peaks are correctly identified by interpretive software, the
table must be verified at least annually with standard materials or organisms with known
characteristics.
REFERENCES
MIC.32822 HPLC Method Validation Phase II

The HPLC method has been validated using known strains of bacteria, including strains
expected to be encountered in routine clinical use.
✓ Record of method validation with appropriate strains
REFERENCES
MIC.32860 HPLC Result Review Phase II

There is a policy for review of HPLC results in conjunction with other laboratory data prior
to reporting results.
NOTE: HPLC is only one tool for microbial identification. When results of HPLC analysis conflict
with growth characteristics, pigmentation, or the results of biochemical or molecular testing,
identification decisions must be based on all the information available.
REFERENCES
MIC.32898 HPLC Analysis - Pure Isolates Phase II

There are procedures to check the purity of cultures used as a source for HPLC analysis.
NOTE: Results of HPLC analysis may be unreliable if mixed cultures are tested. If HPLC is
performed on an isolate from liquid culture and an interpretable chromatogram is obtained, it is
not necessary to await the results of the purity check before reporting results, but a purity check
must still be performed.
REFERENCES
MIC.32936 HPLC Reagent Grade Phase I

Reagents and solvents are of appropriate grade, and solvent purity is assessed when
needed.
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NOTE: Only HPLC grade solvents are recommended for this procedure. Degradation begins
once ultra-pure solvents are opened. Degradation can be slowed by storing solvents in tightly
capped, amber bottles in the dark. Solvent purity verification is suggested when a degradation-
related problem is suspected.
✓ Reagent logs
REFERENCES

There are written procedures for operation and calibration of the HPLC instrument.
NOTE: Basic principles of HPLC analysis require continual monitoring of analysis conditions,
including standard operating procedures, and system calibration. System problems and
corrective actions must be appropriately recorded.
REFERENCES
LABORATORY SAFETY
Laboratory General checklist, to assure that the mycobacteriology laboratory is in compliance. Please elaborate
upon the location and the details of each deficiency in the Inspector's Summation Report.
The following requirements pertain specifically to the mycobacteriology laboratory.
● Mycobacteriology specimen collection, transport and handling policy
● Patient mycobacteriology specimens (sealed, leak proof containers)

● Sealed, safety centrifuge carriers
MIC.33050 Specimen Collection Phase II

All specimens for mycobacterial testing are collected and/or received in sealed leak-proof
containers.
MIC.33100 Centrifuge Safety Phase II

Sealed screw-capped tubes are enclosed in sealed safety centrifuge carriers (i.e. a double
closure system) to minimize aerosol hazards when centrifuging specimens.
REFERENCES
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1) Clinical and Laboratory Standards Institute (CLSI). Clinical Laboratory Safety; Approved Guideline, Third Edition. CLSI document
GP17-A3 [ISBN 1-56238-797-9 (Print); ISBN 1-56238-798-7 (Electronic)]. Clinical and Laboratory Standards Institute, 940 West

The biological safety cabinet meets minimum requirements for mycobacterial work.
NOTE: Exhaust air from a class I or class II biological safety cabinet must be filtered through
high efficiency particulate air (HEPA) filters. Air from Class I and IIB cabinets is hard-ducted to
the outside. Air from Class IIA cabinets may be recirculated within the laboratory if the cabinet is
tested and certified at least every 12 months. It may be exhausted through a dedicated stack that
protects against backflow of air from adverse weather conditions or through the building exhaust
air system in a manner (e.g. thimble connection) that avoids any interference with the air balance
of the biological safety cabinet or building exhaust system.
✓ Written policy defining the types of safety cabinets, filtration systems and exhaust systems
used AND
✓ Records of testing and certification AND
✓ Records of HEPA filters used for filtration of all BSC classes AND
✓ Records of exhaust mechanism OR recirculation, if appropriate
REFERENCES
2) Clinical and Laboratory Standards Institute. Protection of Laboratory Workers From Occupationally Acquired Infections; Approved
th
Guideline. 4 ed. CLSI Document M29-A4. Clinical and Laboratory Standards Institute, Wayne, PA; 2014.
2012.
MYCOLOGY
QUALITY CONTROL
● Sampling of mycology media/stain/reagent QC policies and procedures
● Sampling of mycology QC records
● How do you determine when QC is unacceptable and when corrective actions are
needed?

determine if the steps taken follow the laboratory policy for corrective action
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MEDIA

1. Sterility
NOTE: The laboratory must have records showing that all media used are sterile, able to
support growth, and are appropriately reactive biochemically. This will ordinarily require that the
use. End user quality control must be performed on media not listed on Table 2 of M22-A3 (e.g.
dermatophyte test medium), regardless of the exempt status.
Laboratories receiving media from media suppliers must have a copy of the CLSI/NCCLS
Standard M22-A3 as a reference source. The media supplier must provide records showing
that the quality control activities meet the CLSI/NCCLS Standard M22-A3, or are otherwise
equivalent. The laboratory director may wish to have a signed contractual arrangement with his/
her selected media supplier to cover all expected quality control and documentation thereof.
✓ Written procedure for QC on new lot numbers and shipments of medium AND
✓ Records of media QC AND
applicable
REFERENCES
2) Clinical and Laboratory Standards Institute (CLSI). Principles and Procedures for Detection of Fungi in Clinical Specimens-Direct
Examination and Culture; Approved Guideline. CLSI document M54-A (ISBN 1-56238-857-6 [Print] ISBN 1-56238-858-4 [Electronic]).
Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 2500, Wayne, Pennsylvania 19087-1898 USA, 2012.

appropriate sample of each medium is checked before or concurrent with initial use for
each of the following:
1. Sterility
previous batches
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✓ Written procedure for testing media prepared by the laboratory
Good laboratory practice includes checking all media either at the time of receipt or concurrently with use. This
applies to purchased media as well as media prepared by the laboratory.
MIC.41250 Reference Organisms Phase II

Reference cultures or control organisms are used to check stains and reagents at
appropriate intervals.
REFERENCES
MIC.41270 Nucleic Acid Probe/Exo-antigen QC Phase II

If nucleic acid probes or exo-antigen tests are used for identification of fungi isolated from
culture, appropriate positive and negative controls are tested on each day of use.
✓ Written policy defining QC for nucleic acid probe or exo-antigen tests AND
✓ Records of nucleic acid probe or exo-antigen QC at defined frequency
MIC.41370 Direct Smear Stain QC Phase II

Direct patient specimen stains (e.g. acid fast, PAS, Giemsa, Gomori's methenamine silver,
India ink) are checked with positive and negative controls on each day of patient sample
testing.
NOTE: For certain stains such as GMS and Giemsa, the slide itself serves as the negative
control. Controls for KOH preparations are not required.
✓ Records of stain QC at defined frequency
REFERENCES
1) August, Hindler, Huber, Sewell. Quality control and quality assurance practices. In: Clinical Microbiology, Cumitech 3A. Washington,
DC: American Society for Microbiology, 1990
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7166 [42CFR493. 1256(e)(2); 493.1256(e)(3); 493.1273(a)]
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3709 [42CFR493.1263(a)]

Fluorescent stains (such as calcofluor white) are checked with positive and negative
controls each time of use and records maintained.
REFERENCES
MIC.41400 Lactophenol Cotton Blue QC Phase II

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Lactophenol cotton blue is checked for intended reactivity with a control organism with
each new batch, lot number, and shipment of reagent.
✓ Records of QC at defined frequency
The intent of this series of requirements is to ensure the use of an appropriate variety of media and growth
conditions to isolate the significant pathogens with minimal interference from contaminants.
● Sampling of mycology test procedures/identification schemes
● What tests or procedures are used to identify dimorphic fungi? Filamentous fungi?
● Follow a mycology patient worksheet from receipt and processing to culture

evaluation, identification and reporting
MIC.42025 Cryptococcal Antigen Phase II

If cryptococcal antigen-detection methods are used on CSF, back-up cultures are
performed on positive CSF specimens submitted for diagnosis.
NOTE: It is important to recover the causative organism for precise identification (C. neoformans
vs. C. gattii) and potential susceptibility testing. Back-up cultures of follow-up specimens used for
trending the antigen titer are not required. If culture is not performed onsite by the laboratory, the
laboratory must show evidence that it has been performed in a referral laboratory.
✓ Written policy stating that CSF cultures are performed in conjunction with initial positive
cryptococcal antigen tests OR policy describing testing at another location AND
✓ Records of back-up CSF cultures performed on-site OR records indicating that cultures are
performed at another location
MIC.42050 Selective Media Phase II

Suitable selective media are used for the growth and isolation of dermatophytes and/or
systemic fungi.
✓ Written policy for mycology culture defining the media used for growth and isolation
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MIC.42100 Selective Media Phase II

Media with antimicrobial agents are used to suppress the growth of contaminants.
NOTE: Antimicrobial agents may inhibit some yeasts and the yeast phase of dimorphic
organisms. Both types of media (with and without antimicrobials) should be available and used
when indicated.
✓ Written policy for mycology culture defining the use of media to suppress contaminants
REFERENCES

Incubation temperatures for the growth and isolation of dermatophytes and systemic
fungi are defined and followed under culture conditions.
✓ Temperature records

If cultures are incubated at room temperatures, actual ambient temperature (22 to 26°C) is
recorded daily to determine if proper growth conditions are being maintained.
MIC.42250 Differential Tests Phase II

Procedures for the differentiation and identification of fungi (differential tests) are
adequate for the needs of the laboratory.
NOTE: Laboratories offering full identification must have sufficient procedures to do so. Smaller
laboratories with limited services should have an arrangement with an approved referral
laboratory for back-up and complete identification of mycology specimens.
✓ Written procedure detailing tests performed and identification scheme appropriate for the
extent of testing
REFERENCES
1) Riddle DL, et al. Clinical comparison of the Baxter Microscan yeast identification panel and the Vitek yeast biochemical card. Am J
Clin Pathol. 1994;101:438-442
2) Love GL. Mycology Benchtop Reference Guide: An Illustrated Guide for Commonly Encountered Fungi. Northfield, IL: College of
American Pathologists; 2013.
MIC.42350 Differential Tests Phase II

Differential tests include biochemical tests (e.g. urease, carbohydrate assimilation and/or
fermentation).
MIC.42400 Differential Tests Phase I

Differential tests include slide cultures (when appropriate).
REFERENCES
MIC.42550 Dimorphic Fungi Phase I

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The identification of dimorphic fungal isolates is confirmed by exo-antigen, molecular,

yeast-mold conversion or tissue phase detection tests.
NOTE: Exo-antigen tests, DNA probes, or DNA sequencing are recommended.

REFERENCES
MYCOLOGY SUSCEPTIBILITY TESTING

● Sampling of susceptibility test, QC and reporting policies and procedures
● Sampling of susceptibility QC records
● Susceptibility test set-up (standardized inoculum, pure culture)
● How does your laboratory work with the pharmacy and medical staff to determine
guidelines for reporting of antimicrobial agents?
MIC.42600 Susceptibility Testing QC Phase II

For antifungal susceptibility testing, quality control is performed on each new batch,
lot number and shipment of disks, antimicrobial strips, antimicrobial agents, media,
minimum inhibitory concentration (MIC) panels, and susceptibility testing plates before or
concurrent with initial use with appropriate QC organisms.
✓ Records of new lot susceptibility QC for media, disk, and MIC panels
REFERENCES
1) Department of Health and Human Services, Centers for Medicare and Medicaid Services, Clinical laboratory improvement
amendments of 1988; final rule. Fed Register. 2003(Jan 24):[42CFR493.1263(b)].
MIC.42620 Susceptibility Testing QC Frequency Phase II

For antifungal susceptibility testing by either disk, strip or MIC methods, quality control
organisms are tested with each new lot number and/or shipment and each day the test is
performed thereafter.
NOTE: The frequency of QC testing may be reduced to weekly (and whenever any reagent
component of the test is changed) if the laboratory director approves the use of an individualized
quality control plan (IQCP), including all required elements of IQCP, and the laboratory has
records of satisfactory performance with daily QC tests as suggested by CLSI Standards and
Guidelines (M27, M44, and M38). Please refer to the Individualized Quality Control Plan section
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of the All Common Checklist for the requirements for implementation and ongoing monitoring of
an IQCP.
✓ Records of susceptibility QC results recorded at defined frequency and meeting defined
acceptability criteria
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):[42CFR493.1263(b)(2)].
2) Clinical and Laboratory Standards Institute. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts;
rd
Approved Standard. 3 ed. CLSI Document M27-A3. Clinical and Laboratory Standards Institute, Wayne, PA; 2008.
3) Clinical and Laboratory Standards Institute. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Filamentous
Fungi; Approved Standard. 2
nded. CLSI Document M38-A2. Clinical and Laboratory Standards Institute, Wayne, PA; 2008.
4) Clinical and Laboratory Standards Institute. Method for Antifungal Disk Diffusion Susceptibility Testing of Yeasts; Approved
Guideline. 2
nd ed. CLSI Document M44-A2. Clinical and Laboratory Standards Institute, Wayne, PA; 2009.
MIC.42640 Susceptibility Testing - Pure Cultures Phase II

Only isolated colonies or pure cultures are used for performance of antifungal
susceptibility testing (i.e. susceptibility testing is not performed on mixed cultures).
✓ Written procedure describing the use of isolated colonies or of pure cultures for susceptibility
testing, including the use of purity plates
MIC.42660 Standardized Inoculum Phase II

The inoculum size of both QC strains and test organisms is standardized using a turbidity
standard or another acceptable method.
✓ Written procedure for standardizing susceptibility inoculum
REFERENCES
rd
2) Clinical and Laboratory Standards Institute. Reference Method for Broth Dilution Antifungal Susceptibility Testing of Yeasts; 4th
Informational Supplement. CLSI Document M27-S4. Clinical and Laboratory Standards Institute, Wayne, PA; 2012.
MIC.42680 Susceptibility Testing Endpoint Determination Phase II

For antifungal susceptibility testing, there are written criteria for measuring and
determining the MIC endpoint or zone size.
NOTE: The laboratory may use CLSI criteria, but the use of other validated criteria such as the
FDA or European Committee on Antimicrobial Susceptibility Testing (EUCAST) is acceptable.
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):[42CFR493.1263(b)(1)].
rd
nded. CLSI Document M38-A2. Clinical and Laboratory Standards Institute, Wayne, PA; 2008.
4) Clinical and Laboratory Standards Institute. Method for Antifungal Disk Diffusion Susceptibility Testing of Yeasts; Approved
Guideline. 2
MIC.42700 Antifungal Agents to Test/Report Phase II

There are written policies to ensure that only antifungal agents appropriate for the
organism and body site are routinely tested and reported.
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NOTE: The microbiology department should consult with the medical staff and pharmacy to
develop a list of antifungal agents to be reported for specific organisms isolated from certain
body sites, instead of indiscriminant susceptibility testing and reporting of all fungal isolates or
reporting of all antifungal agents that might be included on a test panel. Isolates from body sites
for which susceptibility might be routinely tested and reported include Candida spp. isolates from
blood cultures.
✓ Patient records showing selection and testing of fungal isolates and reporting of fungal
agents for certain body sites AND
✓ Records of review of antimicrobial reporting policies on an annual basis AND
✓ Proficiency testing susceptibility reporting following policy
REFERENCES
rd
MIC.42720 Inconsistent Antifungal Susceptibility Reports Phase I

There is a written policy to address unusual or inconsistent antifungal testing results.
NOTE: Acceptable results obtained when testing QC organisms do not guarantee accurate
results on patient isolates. Results from testing of patient isolates should be reviewed, and
unusual or inconsistent results should be investigated. Each laboratory should have a policy for
confirming unusual or inconsistent results. For yeasts and molds, the time of endpoint reading
(particularly for the echinocandins) and the effect of trailing growth (particularly for the azoles
and flucytosine) can be significant factors impacting susceptibility results. In some cases, it may
be necessary to repeat susceptibility testing and/or identification procedures to confirm initial
results. This may involve using alternative testing methods or sending the isolate to a referral
laboratory. Options also include retrospectively reviewing individual patient data or cumulative
data for unusual resistance patterns. Some examples of inconsistent antifungal testing results
include:
1. Candida albicans resistant to all azoles
2. Candida spp. susceptible to azoles but resistant to echinocandins
3. Candida albicans resistant to echinocandins
4. Candida krusei susceptible to fluconazole
✓ Records of investigation for unusual or inconsistent results
LABORATORY SAFETY
Laboratory General checklist, to assure that the mycology laboratory is in compliance. Please elaborate upon
The following requirements pertain specifically to the mycology laboratory.

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● Sampling of mycology safety policies and procedures
● Safe work practices (taping of culture plates, procedures performed under BSC)
MIC.43050 Safety Precautions Phase II

If plate culture media is used in mycology, appropriate safety precautions are taken (such
as taping lid to plate on both sides when not in use or other appropriate measures) to
prevent the accidental opening of a plate.
NOTE: Some authorities recommend the transfer of growing colonies from plate to tubed media,
if the former is routinely used for initial inoculation.
✓ Written policy defining safety precautions for handling mycology culture plates
REFERENCES
**REVISED** 08/17/2016
When working with a colony exhibiting mycelial growth, all transfers are performed within
a biologic safety cabinet, and the use of slide culture techniques is limited, whenever
possible, to work with low virulence organisms.
REFERENCES

When preparing teased preparations or "scotch" tape preps, mycelia are always
submerged in liquid medium (such as lactophenol cotton blue).

organisms considered to be highly contagious by airborne routes.
REFERENCES
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The biological safety cabinet (BSC) is certified annually to ensure that filters are
functioning properly and that airflow rates meet specifications.

The BSC meets minimum requirements for mycologic work.
NOTE: Exhaust air from a class I or class II BSC must be filtered through HEPA filters. Air from
Class I and IIB is hard ducted to the outside. Air from Class IIA cabinets may be recirculated
within the laboratory if the cabinet is tested and certified at least every 12 months. It may be
exhausted through a dedicated stack that protects against backflow of air from adverse weather
conditions or through the building exhaust air system in a manner (e.g. thimble connection) that
avoids any interference with the air balance of the BSC or building exhaust system.
used AND
REFERENCES
2012.
PARASITOLOGY
QUALITY CONTROL
● Sampling of parasitology stain/reagent QC policies and procedures
● Sampling of parasitology QC records
● Ocular micrometer calibration procedure and records
● Reference materials (permanent mounts, photomicrographs or atlas available)

● Zinc sulfate solution (tightly-stoppered)
● Ocular micrometer
MIC.51000 Reference Materials Phase I

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Reference materials, such as permanent mounts, photomicrographs, CLSI/NCCLS

documents M15-A and M28-A2, or printed atlases are available at the work bench to assist
with identifications.
REFERENCES
1) NCCLS. Laboratory Diagnosis of Blood-borne Parasitic Diseases; Approved Guideline. NCCLS document M15-A. NCCLS, Wayne,
PA, 2000.
2) Garcia LS, Shimizu R. Diagnostic parasitology: parasitic infections and the compromised host. Lab Med. 1993;24:205-215
3) Ash R, Orihel TC. Atlas of human parasitology, 4th ed. Chicago, IL: American Society of Clinical Pathology, 1996
4) Clinical and Laboratory Standards Institute (CLSI). Procedures for the Recovery and Identification of Parasites From the Intestinal
Tract; Approved Guideline—Second Edition. CLSI document M28-A2 (ISBN 1-56238-572-0). Clinical and Laboratory Standards
Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2005
5) Garcia LS,. Diagnostic medical parasitology. 4th ed. Washington, DC: ASM Press, 2001
6) Colmer-Hamood JA, Fecal microscopy. Artifacts mimicking ova and parasites. Lab Med. 2001;32:80-84
8) Pritt BS. Parasitology Benchtop Reference Guide: An Illustrated Guide for Commonly Encountered Parasites. Northfield, IL: College
of American Pathologists; 2014.
REAGENTS
MIC.51120 Reagents Phase II

If zinc sulfate is used, the solution is stored in a tightly-stoppered bottle and checked for
specific gravity (1.18 for fresh specimens and 1.20 for formalin-fixed specimens) with a
hydrometer whose scale is large enough to differentiate the two values.
✓ Records for specific gravity checks on the zinc sulfate solution
MIC.51160 Permanent Stain QC Phase II

All permanent parasitology stains are checked for intended reactivity with controls or
reference materials at least monthly (or with each test if performed less frequently than
every month).
NOTE: PVA fixative solutions thoroughly mixed with fresh fecal material that has been seeded
with buffy coat leukocytes usually provides reliable controls for permanent stains.
✓ Records of permanent stain QC at defined frequency
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):7167 [42CFR493.1264(c)]
2) Garcia LS, Diagnostic medical parasitology, 4th ed. Washington, DC: ASM Press, 2001
MIC.51170 Special Stain QC Phase II

Stains that are used to detect specific parasites (e.g. acid fast, fluorescent) are checked
with appropriate control organisms each time of use.
✓ Records of special stain QC each time of use
INSTRUMENTS AND EQUIPMENT
MIC.51210 Ocular Micrometer Phase II

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An ocular micrometer is available for determining the size of eggs, larvae, cysts,
trophozoites, and microfilaria or other bloodborne parasites.
REFERENCES
MIC.51220 Calibration/Recalibration - Ocular Micrometer Phase II

The ocular micrometer has been calibrated for the microscope(s) in which it is used and it
is recalibrated each time the eyepieces or objectives are changed.
NOTE: Calibrations can be checked against a micrometer or other objects of known dimensions.
If there are no changes to a particular microscope's optical components, there is no need to
recheck calibration.
✓ Records of initial calibration and recalibration if applicable
REFERENCES

● Sampling of parasitology test procedures
● Follow a parasitology patient worksheet from receipt and processing to identification

and reporting
STOOLS FOR OVA AND PARASITES
MIC.52100 Ova/Parasite Exam Phase II

The microscopic examination of all stools submitted for an ova and parasite (O&P)
examination includes a concentration procedure and a permanent stain.
NOTE: When a stool specimen is submitted fresh, the usual approach would be to perform a
direct wet preparation (looking for motility), a concentration (helminth eggs/larvae/protozoan
cysts), and the permanent stained smear (identification of protozoa missed by concentration
and confirmation of suspect organisms). As a minimum (and certainly if the stool is submitted in
preservatives), the standard O&P examination would include the concentration procedure and
a permanent stained smear. The main point is to ensure that the permanent stained smear is
performed on all stool specimens, regardless of what was or was not seen in the concentration
wet preparation. Often, intestinal protozoa will be seen in the permanent stained smear, but may
be missed in the concentration examination.
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✓ Written procedures for stool for O&P AND
✓ Patient reports/worksheets with concentration and permanent stain results
REFERENCES
1) Garcia LS, Diagnostic medical parasitology, 4th edition. Washington, DC: ASM Press, 2001
2) Clinical and Laboratory Standards Institute (CLSI). Procedures for the Recovery and Identification of Parasites From the Intestinal
Tract; Approved Guideline—Second Edition. CLSI document M28-A2 (ISBN 1-56238-572-0). Clinical and Laboratory Standards
Institute, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA, 2005
MIC.52190 Stool Number/Timing Phase I

There are written policies (developed with clinicians) for the number and/or timing of
collection of stool specimens submitted for routine parasitology testing.
NOTE: Suggestions made by the authors of a 1996 CAP Q-Probes study (Valenstein et al)
include:
1. Accept no more than two or three specimens/patients without prior consultation with
an individual who can explain the limited yield provided by additional specimens
2. Do not accept specimens from inpatients after the fourth hospital day, without prior
consultation
These recommendations are for diagnostic testing. Different policies may apply to tests ordered
for follow-up.
REFERENCES
1) Yannelli B, et al. Yield of stool cultures, ova and parasite tests, and Clostridium difficile determinations in nosocomial diarrhea. Am J
Infect Control. 1988;16:246-249
2) Morris AJ, et al. Application of rejection criteria for stool ovum and parasite examinations. J Clin Microbiol. 1992;30:3213-3216
3) Valenstein P, et al. The use and abuse of routine stool microbiology. A College of American Pathologists Q-probes study of 601
institutions. Arch Pathol Lab Med. 1996;120:206-211
4) Cartwright CP. Utility of multiple-stool-specimen ova and parasite examinations in high-prevalence setting. J Clin Microbiol.
1999;37:2408-2411
BLOOD FILMS FOR MALARIA AND OTHER PARASITES
MIC.52193 Blood Parasite Detection Phase II

The microscopic examination of blood films submitted for detection of blood parasites
allows for detection of parasites responsible for malaria, babesiosis, trypanosomiasis and
filariasis.
REFERENCES
PA, 2000.
MIC.52195 Percentage Parasitemia Reporting Phase I

When blood films are positive for malaria parasites (Plasmodium spp.), the percentage
parasitemia is reported along with the organism identification.
NOTE: It is important to report the percentage of parasitemia when blood films are reviewed and
found to be positive for malaria parasites. Because of the potential for drug resistance in some
of the Plasmodium species, particularly P. falciparum, it is important that every positive smear
be assessed and the parasitemia reported exactly the same way on follow-up specimens as on
the initial specimen. This allows the parasitemia to be followed after therapy has been initiated.
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The parasitemia will usually drop very quickly within the first 24 hours; however, in cases of drug
resistance, the level may not decrease, but actually increase over time.
Although there are currently no requirements for reporting percent parasitemia when blood films
are positive for Babesia species, physicians may ask for these data to monitor the response to
therapy.
✓ Written procedure for performing and reporting parasitemia percentage with identification
REFERENCES
PA, 2000.
2) Garcia LS, Diagnostic Medical Parasitology. Washington, DC, ASM Press, 2001
3) http://www.dpd.cdc.gov/DPDx/HTML/DiagnosticProcedures.htm
MIC.52200 Thick and Thin Films Phase II

Both thick and thin films (routine blood films and/or buffy coat films), or methods of
equivalent sensitivity, are made to provide thorough examination for blood parasites.
REFERENCES
PA, 2000.
2) Thomson S, et al. External quality assessment in the examination of blood films for malarial parasites within Ontario, Canada. Arch
Pathol Lab Med. 2000;124:57-60
MIC.52220 Malaria Stain Procedure Phase I

There are records that malaria stains are washed with a buffer of a pH appropriate for the
stain used (e.g. pH 6.8-7.2 for Giemsa), or the range specified by the manufacturer.
REFERENCES
PA, 2000.
2) Garcia LS, Diagnostic medical parasitology. 4th ed. Washington, DC: ASM Press, 2001
MIC.52260 Slide Review Procedure Phase II

An adequate number of fields are examined under oil immersion using the 100X oil
immersion objective (e.g. 300 fields).
✓ Written procedure defining criteria for assessment of malaria slides including objective and
number of fields examined
REFERENCES
PA, 2000.
LABORATORY SAFETY
Laboratory General checklist, to assure that the parasitology laboratory is in compliance. Please elaborate upon
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The following requirements pertain specifically to the parasitology laboratory.
● Formalin monitoring procedure and records of monitoring
**REVISED** 08/21/2017
MIC.53050 Formaldehyde and Xylene Safety Phase II
Formaldehyde and xylene vapor concentrations are maintained below the following
maxima, expressed as parts per million.
NOTE: Formaldehyde and xylene vapor concentrations must be monitored in all areas where
these reagents are used. Initial monitoring involves identifying all employees who may be
exposed at or above the action level or at or above the STEL and accurately determining
the exposure of each employee identified. Once an initial monitoring procedure has been
performed, further periodic formaldehyde monitoring is mandated at least every six months
if the initial monitoring result equals or exceeds 0.5 ppm (8 hr time-weighted exposure, the
“action level”) or at least once per year if the results exceed the short term exposure limit (STEL)
2.0 ppm. The laboratory may discontinue periodic formaldehyde monitoring if results from two
consecutive sampling periods taken at least seven days apart show that employee exposure
is below the action level and the short-term exposure limit, and 1) no change has occurred in
production, equipment, process or personnel or control measures that may result in new or
additional exposure to formaldehyde, and 2) there have been no reports of conditions that may
be associated with formaldehyde exposure.
Formaldehyde monitoring must be repeated any time there is a change in production, equipment,
process, personnel, or control measures which may result in new or additional exposure to
formaldehyde for an employee involved in the activity. If any personnel report signs or symptoms
of respiratory or dermal conditions associated with formaldehyde exposure, the laboratory must
promptly monitor the affected person's exposure.
Xylene must be monitored initially, but there is no requirement for periodic monitoring of xylene.
Repeat monitoring should be considered when there is a change in production, equipment,
process, personnel, or control measures likely to increase exposure levels.
8 hr Time- Action Level (8 hr 15 min Short-Term

Weighted Exposure Time-Weighted Average Exposure
Limit in ppm Exposure) in ppm Limit (STEL) in ppm
Formaldehyde 0.75 0.5 2.0
Xylene 100 150
✓ Written safety policy for formalin and xylene safety including action limits, criteria for
discontinuation of monitoring and criteria for resumption of monitoring AND
✓ Record of initial formalin and xylene monitoring and repeat monitoring when indicated AND
✓ Records of corrective action when exposure limits are exceeded
REFERENCES
1) Montanaro A. Formaldehyde in the workplace and in the home. Exploring its clinical toxicology. Lab Med. 1996;27:752-757
2) Goris JA. Minimizing the toxic effects of formaldehyde. Lab Med. 1997;29:39-42
3) Occupational Safety and Health Administration, 1998(Jul 1) [29CFR1910.1048]
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MIC.53150 Ether Safety Phase II

If a procedure uses ether, the diethyl ether is stored on open shelves in a well ventilated
room using the smallest can feasible (as shipped by manufacturer).
NOTE: The use of concentration techniques other than those requiring the use of ether is
recommended.
VIROLOGY
QUALITY CONTROL
● Sampling of virology test procedures
● Sampling of virology cell culture system and reagent QC policies and procedures
● Sampling of virology QC records and cell line checks
● How are cell lines checked for contamination?
● Follow a virology patient worksheet from receipt and processing to identification and
reporting
REAGENTS
MIC.61000 Breakage Phase I

The laboratory has records that each shipment of commercial cell culture tubes, flasks,
shell vials or cluster trays is examined for breakage.
REFERENCES
MIC.61050 Acceptance of Materials Phase II

There are written policies for the acceptance and rejection of cell culture tubes, flasks,
shell vials or cluster trays used for virus isolation.
NOTE: Cell cultures should be observed microscopically to confirm that the cells are attached
to the substratum, the confluency of the monolayer is appropriate for the method and cell
line (75%-90%) and cell appearance is typical. Confluent or overgrown monolayers may
obscure viral cytopathic effect (CPE) in tube or flask cell cultures and can adversely affect the
recovery of some more fastidious viruses, such as RSV. The cell culture media should be free
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of contamination (clear) and should be near a neutral ph (salmon pink in color). Cell culture
tubes, flasks, vials or plates not meeting the expected criteria should be observed or rejected,
depending on the observation and laboratory policies.
REFERENCES
**NEW** 08/21/2017
MIC.61060 Cell Culture System Growth Checks Phase II
The cell culture system is checked for the ability to support growth.
NOTE: For systems using shell vials and/or co-cultivated cell lines that detect multiple viruses,
laboratories can evaluate both the ability of the cell lines to support the growth of the virus(es)
and the reactivity of the detection reagents (MIC.61370) by rotating growth controls that include
the viruses tested for which the test is performed. By using this method over the course of a
week, the cell culture system lot and detection reagents will be checked for all viruses. While
rotation of viral culture controls for all viral targets is desirable, certain viruses may be difficult
to maintain by serial propagation (for example varicella zoster virus). In these cases, integrity of
the cell culture system may be demonstrated through the use of other, more easily propagated
viruses.
✓ Written procedure for cell culture system QC AND
✓ Records of quality control
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24): [42CFR493.1265(a)].
**REVISED** 08/21/2017
MIC.61140 Cell Culture Media and Diluent Checks Phase II
Cell culture media and diluents are checked for sterility and pH.
NOTE: Entering the media to remove aliquots for refeeding, etc. does not generate the need
for repeat sterility testing. It is satisfactory to test either the individual components or the final
product.
✓ Written procedure for cell culture media QC AND
✓ Records of media sterility and pH QC
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):[42CFR493.1265(a)].
MIC.61150 Cell Line QC Phase II

Continuous cell lines are checked for mycoplasma contamination.
NOTE: An alternative method to culturing for mycoplasma is the monitoring of a negative,

uninoculated control. If cell lines are provided by a commercial vendor, documentation of
mycoplasma screening by the vendor is acceptable. Vendor records must be retained by the
laboratory.
REFERENCES
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2) Lennette DA. General Principles for Laboratory Diagnosis for Viral, Rickettsial, and Chlamydial Infections. In: Lennette EH, Lennette,
DA, Lennette ET, editors. Diagnostic Procedures for Viral, Rickettsial, and Chlamydial Infections. Seventh Edition ed. Washington:
American Public Health Association; 1995 p. 3-25
MIC.61155 Cell Line QC Phase II

Continuous cell lines are checked for endogenous viral contamination.
NOTE: Upon receipt in the laboratory and during the period of use, cell lines must be monitored
for the presence of endogenous contamination due to viruses such as foamy virus and monkey
virus.
Endogenous viral contamination must be recorded. Individual laboratories may decide to reject
all cell lines with contamination. Alternatively, cell lines may be monitored to determine if the
contamination will prohibit the isolation and identification of patient viral isolates. Endogenously
contaminated cell cultures can be evaluated by considering conditions such as the degree
(percent) of contamination of the monolayer, the specificity of the contamination CPE, the effect
of the contamination on the quality of the culture system, and the condition of the culture cell to
support virus growth. If such contamination will affect the ability to recover patient isolates, cell
lines must be rejected.
REFERENCES
MIC.61160 Media QC Phase II

Animal sera used for cell growth media are checked for absence of toxicity to cells.
✓ Written procedure for checks of animal sera for toxicity AND
✓ Records of animal sera checks
MIC.61180 Cell Line Availability Phase II

The laboratory has the appropriate minimal cell line(s) available for all types of specimens
tested and for all viruses reported by the laboratory.
NOTE: The following is a suggested list of cell lines for the intended purpose:
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ORGANISM CELL LINE

Chlamydia McCoy or Buffalo Green Monkey Kidney
Herpes simplex HDF, Primary or First Pass Rabbit Kidney, MRC-5, or A549, CV-1/
MRC-5, transgenic BHK
Varicella zoster HDF, Primary Monkey Kidney, CV-1/MRC-5
Influenza Primary Monkey Kidney or MDCK, Mink lung /A549
Parainfluenza Primary Monkey Kidney, Mink lung/A549
RSV HEp-2 or Primary Monkey Kidney, Mink lung/A549
Enteroviruses Primary Monkey Kidney, A549, HDF, MRC-5, GGMK+Daf/A549
Adenoviruses HEp-2, Human Embryonic Kidney, A549, Mink lung/A549, MDCK/
A549
Human metapneumovirus LLC-MK2, Mink lung/A549
Cytomegalovirus HDF, MRC-5
REFERENCES
1) McCarter YS, Robinson A. Comparison of MRC-5 and primary rabbit kidney cells for the detection of Herpes simplex virus. Arch
Pathol Lab Med. 1997;121:122-124
2) McCarter YS, Ratkiewicz IN. Comparison of virus culture and direct immunofluorescent staining of cytocentrifuged virus transport
medium for detection of Varicella-Zoster virus in skin lesions. Am J Clin Pathol. 1998;109:631-633
**REVISED** 08/17/2016
MIC.61210 Incubation Time Phase II
Viral cultures are incubated for a sufficient time to recover the viruses for which service is
offered.
NOTE: The following is a suggested list of minimum incubation times for the intended purpose:
ORGANISM INCUBATION TIME

Herpes simplex (genital) 5 days
Herpes simplex (other) 7 days
Respiratory viruses 10 days - 14 days
Other viruses 14 to 28 days
✓ Written procedure indicating length of incubation for each virus cultured
REFERENCES
2) Leland DS, Ginocchio CC. Role of cell culture for virus detection in the age of technology. Clin Microbiol Rev 2007 January;
20(1):49-78
3) Landry ML, Hsiung GD. Primary Isolation of Viruses. In: Specter S, Hodinka RL, Young SA, editors. Clinical Virology Manual. Third
Edition ed. Washington: ASM Press; 2000. p. 27-42
4) Leland DS, Emanuel D. Laboratory diagnosis of viral infections of the lung. Semin Respir Infect 1995 December;10(4):189-98
MIC.61300 Work Records Phase II

Records are kept of cell types, passage number, source, and media that are used for
specimen testing, virus culture or propagation and maintenance of control organisms.
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**REVISED** 08/17/2016
MIC.61310 Cytopathic Effect Phase II
Inoculated cultures are checked for cytopathic effect in a fashion that optimizes the time
to detection of viral pathogens.
NOTE: Primary cultures must be checked at least every other working day for cytopathic effect at
least for the first two weeks of incubation.
✓ Written policy defining the frequency of CPE checks AND
✓ Work records reflecting CPE examination at defined frequency
MIC.61320 Media QC Phase II

Uninoculated cell monolayers or monolayers that have been inoculated with sterile
material are available for comparison with cultures of clinical material.
NOTE: Uninoculated cell culture controls must be included on each inoculation day of cell culture
tubes in order to detect non-specific degeneration; or to detect extraneous infection of the cell
culture with endogenous viral agents capable of producing cytopathic effects.
✓ Records of uninnoculated/sterile cell monolayer checks documented at defined frequency
REFERENCES
MIC.61325 Unusual Cytopathic Effect Phase I

There are written policies for the handling of cell cultures with unusual cytopathic effect.
NOTE: Unusual CPE can be detected in cell culture from samples positive for viruses not
commonly detected by the laboratory (i.e. monkey pox, vaccina, variola, etc.) Policies should
include the extent of further manipulation of the cell culture and notification of the appropriate
regulatory agency (ex. Department of Health, CDC).
MIC.61340 Standardized Red Cell Suspensions Phase I

Red cell suspensions that are used for quantitative serologic procedures are standardized
(photometrically or with some other equivalent procedure).
✓ Written procedure for standardization of red cell suspensions
MIC.61350 Agglutination/Lysis Criteria Phase I

Criteria for degrees of agglutination and lysis are defined for quantitative serologic
procedures.
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MIC.61360 Work Records Phase I

Worksheets and/or records indicate actual titers, when known, of reagents and control
sera.
**REVISED** 08/21/2017
MIC.61370 Serologic Reaction Controls Phase II
Reactive and nonreactive controls are analyzed in serologic reactions for detection of
antigens.
NOTE: Positive and negative controls must be run daily for immunofluorescent and
immunochromatic testing when using pool reagents and for virus specific reagents, if performed.
✓ Written procedure for serological QC AND
✓ Records of serological QC results
MIC.61380 Reagent Verification Phase II

Each new lot and shipment of reagents that detect multiple viruses are verified for each
individual virus component prior to patient testing
NOTE: A pool reagent cannot be verified using only a pool control, as the reactivity of each virus
specific component cannot be individually assessed. After initial verification, pool controls can be
used for daily quality control of the pool reagent.
✓ Records of IF reagent verification, as applicable
REFERENCES
TESTS AND PROCEDURES

● Sampling of virology test procedures
● Sampling of virology patient reports
MIC.62400 Order Information Phase I

For viral screening tests by direct antigen detection (direct immunofluorescence or EIA),
rapid cell culture or molecular methods, reports and test order information indicates the
specific viruses sought/detected by the assay.
NOTE: For example, if the rapid cell culture method is used to detect seven different respiratory
viruses, then the report must specifically indicate which viruses are included in the screening.
While the cell lines in use may permit the growth of other viruses, such as enterovirus, these
need not be specifically enumerated in the report, unless detected in a given sample.
MIC.62500 Isolation Policies Phase I

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There are written policies for viral testing based upon such criteria as specimen source,
diagnosis, suspected virus(es) and season.
NOTE: Testing algorithms can vary depending on specimen type, virus(es) suspected, immune
status of the patient, and season. For example, routine rapid EIA testing for influenza is not
recommended outside of the respiratory virus season due to low specificity.
REFERENCES
1) Leland DS, Ginocchio CC. Role of cell culture for virus detection in the age of technology. Clin Microbiol Rev 2007 January;
20(1):49-78
MIC.62550 CMV Antigenemia Phase I

There are written policies for the acceptance and rejection of samples for CMV
antigenemia testing.
NOTE: Policies must be in place to deal with suboptimal specimens, such as those whose
receipt in the laboratory exceeds the time frame for optimal test sensitivity, or those samples
with low cellularity. If it is not possible to recollect a sample, and such specimens are tested,
results must be accompanied by a comment noting the potential reduction in test reliability due to
inappropriate sample storage and/or delay in processing.
REFERENCES
1) Boeckh M, Boivin G. Quantitation of cytomegalovirus: methodologic aspects and clinical applications. Clin Microbiol Rev 1998
July;11(3):533-54
MIC.62560 Tzanck Test Phase I

Slides for the Tzanck test are of sufficient quality for diagnosis.
NOTE: Slides must be of adequate technical quality to be diagnostically useful. The laboratory
should have access to a photographic atlas appropriate to the diagnostic purpose and method
(e.g. Papanicolaou or Giemsa) in use.
LABORATORY SAFETY
Laboratory General checklist, to assure that the virology laboratory is in compliance. Please elaborate upon the
location and the details of each deficiency in the Inspector's Summation Report.
The following requirements pertain specifically to the virology laboratory.
● Sampling of virology specimen handling and processing policies

organisms considered to be highly contagious by airborne routes.
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REFERENCES
2) Biological Safety, Principles and Practices, 4th ed. Fleming DO, Hunt DL, Ed. (ISBN 978-1-55581-339-0) ASM Press; Washington
DC., 2006
Guideline. 4
th ed. CLSI Document M29-A4. Clinical and Laboratory Standards Institute, Wayne, PA; 2014.

The BSC is certified annually to ensure that filters are functioning properly and that
airflow rates meet specifications.

The BSC meets minimum requirements for virology work.
NOTE: Exhaust air from a class I or class II BSC must be filtered through HEPA filters. Air from
Class I and IIB is hard ducted to the outside. Air from Class IIA cabinets may be recirculated
within the laboratory if the cabinet is tested and certified at least every 12 months. It may be
exhausted through a dedicated stack that protects against backflow of air from adverse weather
conditions or through the building exhaust air system in a manner (e.g. thimble connection) that
avoids any interference with the air balance of the BSC or building exhaust system.
used AND
REFERENCES
2) Biological Safety, Principles and Practices, 4th ed. Fleming DO, Hunt DL, Ed. (ISBN 978-1-55581-339-0) ASM Press; Washington
DC., 2006
2012.
MIC.63200 Specimen Handling/Processing Phase II

There are written policies for the safe handling and processing of virology specimens.
✓ Written policies for safe handling/processing of specimens
REFERENCES
Guideline. 4
th ed. CLSI Document M29-A4. Clinical and Laboratory Standards Institute, Wayne, PA; 2014.
**REVISED** 08/17/2016
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There are written policies for the safe handling and processing of samples that are
suspected to contain highly infectious emerging pathogens.
NOTE: Laboratories should review State and Federal guidelines for the handling of samples from
patients suspected to have high risk pathogens such as avian influenza, MERS coronavirus, or
SARS coronavirus.
REFERENCES
MIC.63250 Hazardous Waste Disposal Phase II

Specimens and used media are disinfected, sterilized or contained in a manner to
minimize the hazard of an accident during transportation to a remote autoclave or
incinerator.
✓ Written procedure for the handling and disposal of microbiology waste
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Clinical Laboratory Waste Management; Approved Guideline—Third Edition. CLSI
document GP05-A3 (ISBN 1-56238-744-8). Clinical and Laboratory Standards Institute, 940 West Valley Road, Suite 1400, Wayne,
Pennsylvania 19087-1898, USA 2011.
MOLECULAR MICROBIOLOGY
This checklist section does not apply to tests using direct non-amplified nucleic acid probes to identify
organisms from a positive culture. Such tests may be inspected with the Mycobacteriology, Mycology or other
appropriate section of this checklist.
This checklist section applies to all other molecular microbiology tests, including FDA-cleared/approved tests,
as well as tests not cleared/approved by the FDA (including FDA-cleared/approved tests modified by the
laboratory, and laboratory-developed tests). A database of FDA-cleared/approved tests can be found at http://
www.accessdata.fda.gov/scripts/cdrh/cfdocs/cfCLIA/Search.cfm
When specimens are referred to outside referral laboratories for sequence analysis or other testing, such
laboratories must meet the requirements in GEN.41350 and other applicable requirements in the "Reporting of
Results" section of the Laboratory General checklist.
Laboratories that use this section of the checklist must also comply with all applicable requirements included in
the General section of the Microbiology checklist.
QUALITY MANAGEMENT
● Sampling of QM statistics/turnaround time data
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● What is your course of action when monitored statistics increase above the expected
positive rate?
MIC.63252 Statistics Phase I

When appropriate, statistics (e.g. percentage of results that are positive for Chlamydia
trachomatis and/or Neisseria gonorrhoeae) are maintained and monitored.
NOTE: An increase above the expected positive rate within a run or over multiple runs should
prompt investigation for potential false positive results.
✓ Written procedure for calculating statistics including thresholds AND
✓ Records of statistical data, evaluation and corrective action if indicated
MIC.63256 Turnaround Times Phase I

There is evidence that the laboratory monitors sample turnaround times and that they are
appropriate for the intended purpose of the test.
NOTE: There are certain clinical situations in which rapid completion is essential. An example is
detection of HSV in CSF.
✓ Written policy defining turnaround time and mechanism for monitoring AND
✓ Records showing that times defined in the policy are routinely met
SPECIMEN HANDLING & PROCESSING
● Sampling of molecular microbiology specimen handling and processing policies and
procedures
● Patient specimens/aliquots (storage, labeling)
● What is your process for identification of patient specimens and aliquots?
MIC.63318 Specimen Handling Procedures Phase II

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There are written procedures to prevent specimen loss, alteration, or contamination

during collection, transport, processing and storage.
NOTE: Specimen collection, processing and storage must follow manufacturer's instructions. For
example, for amplified molecular testing using liquid based cervical cytology (LBC) specimens,
there must be a procedure in place to ensure absence of cross contamination of samples during
the LBC processing steps; alternatively, an aliquot can be removed for amplified molecular
testing prior to LBC processing.
MIC.63322 Specimen Aliquots Phase II

If aliquoting of specimens is performed, there is a written procedure that prevents cross-
contamination of the aliquot containers.
NOTE: Although in some cases it may be appropriate to aliquot a specimen, the laboratory must
have a policy that no aliquot is ever returned to the original container.
MIC.63324 Residual Samples Phase II

If residual samples are used for amplification-based testing, policies and procedures
ensure absence of cross-contamination of samples.
NOTE: An example of a residual sample is a liquid based cytology sample that is tested post-
cytologic processing using amplified C. trachomatis or N. gonorrhoeae tests.
MIC.63327 Derivative Material Identification Phase II

There is a system to positively identify derivative material (e.g. nucleic acid extracts) from
patient specimens from nucleic acid extraction through all phases of subsequent testing
and storage.
MIC.63328 Specimen Processing/Storage Phase II

Patient samples are processed promptly or stored appropriately to minimize degradation
of nucleic acids.
NOTE: Frost-free freezers may not be used to store patient samples unless freezer temperature
is monitored by a continuous monitoring system, or a maximum/minimum thermometer.
✓ Written procedure for processing and storage of specimens
REFERENCES
1) Farkas DH, Kaul KL, Wiedbrauk DL, et al. Specimen Collection and Storage for Diagnostic Molecular Pathology Investigation. Arch
Pathol Lab Med. 1996;120:591-596
2) Kiechle FL, Kaul KL, Farkas DH. Mitochondrial Disorders: Methods and Specimen Selection for Diagnostic Molecular Pathology.
Arch Pathol Lab Med. 1996;120:597-603
3) Farkas DH, Drevon AM, Kiechle FL, et al. Specimen Stability for DNA-based Diagnostic Testing. Diag Molec Pathol.
1996;5(4):227-235
ASSAY VALIDATION AND VERIFICATION
Additional requirements and details for validation and verification of methods are found in the "Test Method
Validation and Verification" and "Method Performance Specifications - Nonwaived Test" sections of the All
Common Checklist.
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● Sampling of assay verification and validation studies
● How does your laboratory verify or validate assay performance prior to test
implementation?
● How do you ensure that the modified FDA-cleared/approved test exhibits equal or
superior performance?
**REVISED** 08/21/2017
MIC.64760 Validation or Verification Study Phase II
There are records that the laboratory has performed a validation or verification study prior
to reporting patient results.
NOTE: Refer to the section “Test Method Validation and Verification” in the All Common
Checklist for additional details.
**REVISED** 08/21/2017
MIC.64770 Validation Studies - Specimen Type/Collection Phase II
If the laboratory tests specimen types or uses collection devices other than those listed
in the package insert, the laboratory performs validation studies to document adequate
performance of the test.
NOTE: If the use of an alternative specimen type or collection device requires any part of the
test procedure to be modified (including, for example, a preprocessing step), the test must be
validated.
Results from tests performed on specimen types not listed in the package insert may be reported
without complete validation only if the specimen type is encountered rarely, precluding an
adequate number for validation studies. Under these circumstances, the test report must include
a disclaimer stating that the specimen type has not been validated
MIC.64790 Validation of Specimen Pooling Phase II

If the laboratory chooses to pool specimens for tests performed using test systems
that have not been FDA-cleared/approved for that purpose (e.g. Chlamydia trachomatis/
Neisseria gonorrhoeae NAAT on pooled urine specimens), the testing procedure
for pooled specimens must be validated, including limit of detection (sensitivity),
reproducibility, and accuracy (method comparison).
NOTE: As part of the method comparison, the results for pooled specimens must be compared
to the single (non-pooled) results using an adequate number of clinical specimens covering the
entire range of organism concentration seen in clinical specimens (i.e. low and high positive
specimens).
Any clinical claim regarding the efficacy of pooling must be validated (see COM.40640).
REFERENCES
1) Centers for Disease Control and Prevention. Screening Tests to Detect Chlamydia trachomatis and Neisseria gonorrhoeae
Infections—2002. MMWR 2002;51(No. RR-15).
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MIC.64956 Modified FDA-Cleared/Approved Assay Phase II

If the laboratory modifies an FDA-cleared/approved assay, the modified procedure has
been validated to yield equivalent or superior performance.
✓ Records of validation studies for modified FDA-cleared/approved assays
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Fluorescence In Situ Hybridization Methods for Clinical Laboratories; Approved
Guideline—Second Edition. CLSI document MM07-A2 (ISBN 1-56238-885-1] Clinical and Laboratory Standards Institute, 940 West
2) Lawrence Jennings, Vivianna M. Van Deerlin, Margaret L. Gulley (2009) Recommended Principles and Practices for Validating
Clinical Molecular Pathology Tests. Archives of Pathology & Laboratory Medicine: Vol. 133, No. 5, pp. 743-755
amendments of 1988, final rule. Fed Register. 2003(Jan 24): [42CFR493.1253](b)(2)]
**REVISED** 08/21/2017
MIC.64960 Validation or Verification Studies - Specimen Selection Phase II
Validation or verification studies were performed with an adequate number and
representative (reasonable) distribution of samples for each type of specimen (e.g. blood,
fresh/frozen tissue, paraffin-embedded tissue).
NOTE: Validation or verification studies must include specimens representing each strain
or genotype, as appropriate, and an adequate number of positive and negative specimens
representing the specimen types used in the assay (e.g. plasma, blood, CSF). Specimens may
be spiked (suspensions of target added to appropriate matrix), if patient specimens are not
available or inadequate in number across the dynamic range of a quantitative assay.
For qualitative tests this includes comparison of positive and negative test results to a
comparable test method. For quantitative tests, the manufacturer's limit of detection, linearity,
reportable range and precision must be validated or verified by the laboratory, as well as
a comparison of patient test results across the reportable range of the test. Specimens for
the validation or verification study can include quantitative external control material, cultured
organisms (quantified) and proficiency testing material, and must include patient specimens.
Refer to the section "Test Method Validation and Verification" in the All Common Checklist for
additional details.
✓ Records of validation and verification studies
**REVISED** 08/21/2017
MIC.64968 Validation Study Comparison - Laboratory-Developed Test Phase II
For laboratory-developed tests, the results of each validation study were compared
to another valid test method or to results of a specimen exchange with a laboratory
performing the same type of test in a similar fashion.
NOTE: There may not be a closely related test to be used for comparison. In such cases, the test
performance (sensitivity and specificity) should be assessed in relation to the patient's clinical
diagnosis and in addition assessed by exchanges of specimens with a laboratory that performs
the test in a similar fashion.
✓ Records of comparison and evaluation of each validation study to another test method OR
records of comparison using specimen exchange with another laboratory
MIC.64975 Modified Cut-Off Phase II

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If the laboratory has modified the manufacturer’s cut off-value for a positive result, the
new cut-off value has been validated.
✓ Records of cut-off validation when different cut-off values are utilized
QUANTITATIVE ASSAYS: CALIBRATION & STANDARDS
This section of the checklist only applies to quantitative tests for which appropriate external materials exist.
This introduction discusses the processes of calibration, calibration verification, and analytic measurement
range (AMR) verification.
DEFINITIONS:
CALIBRATION is the set of operations that establish, under specified conditions, the relationship between
reagent system/instrument response and the corresponding concentration/activity values of an analyte.
Calibration procedures are typically specified in the manufacturer's instructions, but may also be established by
the laboratory.
CALIBRATION VERIFICATION denotes the process of confirming that the current calibration settings for each
analyte remain valid for a test system. If calibration verification confirms that the current calibration settings
for each analyte are valid, it is not necessary to perform a complete calibration or recalibration of the test
system. Each laboratory must define limits for accepting or rejecting tests of calibration verification. Calibration
verification can be accomplished in several ways. If the manufacturer provides a calibration validation or
verification process, it should be followed. Other techniques include (1) assay of the current method calibration
materials as unknown specimens, and determination that the correct target values are recovered, and (2) assay
of matrix-appropriate materials with target values that are specific for the test system.
MATERIALS SUITABLE FOR CALIBRATION VERIFICATION

Materials for calibration verification must have a matrix appropriate for the clinical specimens assayed by that
method, and target values appropriate for the measurement system. Suitable materials may include, but are not
limited to:
1. Calibrators used to calibrate the analytical measurement system
2. Materials provided by the analytical measurement system vendor for the purpose of calibration
verification
3. Previously tested unaltered patient/client specimens
4. Primary or secondary standards or reference materials with matrix characteristics and target values
appropriate for the method,
5. Proficiency testing material or proficiency testing validated material with matrix characteristics and
target values appropriate for the method
In general, routine control materials are not suitable for calibration verification, except in situations where the
material is specifically designated by the method manufacturer as suitable for verification of the method's
calibration process.
ANALYTICAL MEASUREMENT RANGE (AMR) is the range of analyte values that a method can directly
measure on the specimen without any dilution, concentration, or other pretreatment not part of the usual assay
process.
AMR VERIFICATION is the process of confirming that the assay system will correctly recover the concentration
or activity of the analyte over the AMR. The materials used for verification must be known to have matrix
characteristics appropriate for the method. The matrix of the sample (i.e. the environment in which the analyte
is suspended or dissolved) may influence the measurement of the analyte. The method manufacturer may
recommend suitable materials. The test specimens must have analyte values that, at a minimum, are near the
low, midpoint, and high values of the AMR. Specimen target values can be established by comparison with
peer group values for reference materials, by assignment of reference or comparative method values, and by
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dilution or admixture ratios of one or more specimens with known values. Each laboratory must define limits for
accepting or rejecting verification tests of the AMR.
MATERIALS SUITABLE FOR AMR VERIFICATION

Materials for AMR verification should have a matrix appropriate for the clinical specimens assayed by that
method, and target values appropriate for the measurement system. Materials may include, but are not limited
to:
1. Linearity material of appropriate matrix, e.g. CAP Survey-based or other suitable linearity
verification material
2. Proficiency testing survey material or proficiency testing survey-validated material
3. Previously tested patient/client specimens, unaltered
4. Previously tested patient/client specimens, altered by admixture with other specimens, dilution,
spiking in known amounts of an analyte, or other technique
5. Primary or secondary standards or reference materials with matrix characteristics and target values
appropriate for the method
6. Calibrators used to calibrate the analytic measurement system
7. Control materials, if they adequately span the AMR.
RECALIBRATION/CALIBRATION VERIFICATION and AMR VERIFICATION INTERVAL:

Recalibration or calibration verification, and AMR verification must be performed at least once every six months.
Successful calibration verification certifies that the calibration is still valid; unsuccessful calibration verification
requires remedial action, which usually includes recalibration. The performance of recalibration or a calibration
verification procedure resets the calendar to a new maximum six-month interval before the next required
reassessment. Test systems that are recalibrated more frequently than every six months do not require a
separate calibration verification procedure.
In addition to the every six-month requirement, laboratories must perform recalibration or calibration verification
and AMR verification at changes in major system components and at changes of lots of chemically or physically
active reagents unless the laboratory can demonstrate that changing reagent lot numbers does not affect the
range used to report patient/client test results. The director should determine what constitutes a verification of
the AMR. Manufacturers' instructions should be followed.
The laboratory should establish other criteria, as appropriate, for recalibration/calibration verification. These
include but are not limited to failure of quality control to meet established criteria, and major maintenance or
service to the instrument.
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● Sampling of calibration and AMR policies and procedures
● Sampling of calibration records
● Sampling of AMR verification records
● Sampling of calibration materials (labeling, storage, quality)
● What is your course of action if calibration is unacceptable?

● When was the last time you performed a calibration procedure and how did you verify
the calibration?
● What is your course of action when results fall outside the AMR?
● Further evaluate the responses, corrective actions, and resolutions for unacceptable
calibration, unacceptable calibration verification, and results outside the AMR
MIC.65100 Calibration Procedures Phase II

Calibration procedures for each test system are appropriate, and the calibration records
are reviewed for acceptability.
REFERENCES
1) Department of Health and Human Services, Centers for Medicare & Medicaid Services. Clinical laboratory improvement amendments
of 1988; final rule. Fed Register. 1992(Feb 28):7165 [42CFR493.1217]
2) Department of Health and Human Services, Centers for Medicare & Medicaid Services. Medicare, Medicaid and CLIA Programs;
Laboratory Requirements Relating to Quality Systems and Certain Personnel Qualifications; final rule. Fed Register. 2003(Jan
24):3707 [42CFR493.1255]
3) Kroll MH, Emancipator K. A theoretical evaluation of linearity. Clin Chem. 1993;39:405-413
4) rd
Clinical and Laboratory Standards Institute. Evaluation of Matrix Effects; Approved Guideline. 3 ed. CLSI Document EP14-A3.
Clinical and Laboratory Standards Institute, Wayne, PA; 2014
5) Miller WG. “Quality control.” Professional Practice in Clinical Chemistry: A Companion Text, ed. DR Dufour. Washington, DC: AACC
Press, 1999:12-1 to 12-22
6) Kroll MH, et al. Evaluation of the extent of non linearity in reportable range studies. Arch Pathol Lab Med. 2000;124:1331-1338
7) Miller WG. Quality Control In: Henry's Clinical Diagnosis and Management by Laboratory Methods, 21st Edition, ed. by McPherson
RA and Pincus MR, Saunders Elsevier 2007, p99-111
**REVISED** 08/21/2017
MIC.65120 Test Calibration - FDA-Cleared/Approved Tests Phase II
For FDA-cleared/approved quantitative tests, test calibration is performed according to
the manufacturer’s specifications.
NOTE: Calibrators must be run following the manufacturer’s recommendations. Some systems
may use electronic calibration data.
✓ Records of calibration
**REVISED** 08/21/2017
MIC.65130 Calibration Materials - LDT and Modified FDA-Cleared/Approved Tests Phase II
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High quality materials with test system and matrix-appropriate target values are used for
calibration and calibration verification for laboratory-developed (LDT) and modified FDA-
cleared/approved tests whenever possible.
NOTE: If a different matrix is used for recalibration of subsequent (different) reagent lots, its
equivalence to the test sample matrix must be established.
For example, if multiple specimen types are tested in a quantitative test, the test calibration must
encompass the range for all expected values for each specimen type.
✓ Written policy defining the use of appropriate calibration/calibration verification materials
REFERENCES
1) Clinical and Laboratory Standards Institute. Evaluation of Matrix Effects; Approved Guideline. 3rd ed. CLSI Document EP14-A3.
Clinical and Laboratory Standards Institute, Wayne, PA; 2014
amendments of 1988, final rule. Fed Register. 2003(Jan 24): [42CFR493.1255]
**REVISED** 08/21/2017
MIC.65140 Calibration Materials - Laboratory Developed Tests Phase II
The quality of all calibration materials used for laboratory-developed tests is evaluated
and recorded.
NOTE: Commercial standards used to prepare calibrators require certificates of analysis. The
laboratory must evaluate the accuracy of a new lot of calibrators by checking the new lot against
the current lot.
REFERENCES
amendments of 1988, final rule. Fed Register. 2003(Jan 24): [42CFR493.1255]
MIC.65150 Recalibration Phase II

The system is recalibrated when calibration verification fails to meet the established
written criteria of the laboratory.
NOTE: An indication of a potential calibration failure would be external or kit controls with values
that repeatedly fall outside of the established control range.
✓ Written policy defining criteria for recalibration AND
✓ Records of recalibration, if calibration or calibration verification has failed
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3707[42CFR493.1255(b)(3)].
MIC.65160 AMR Verification Criteria Phase II

There is a written policy for the verification of the analytical measurement range and
records maintained.
NOTE: The AMR must be verified at least every six months after a method is initially placed in
service and if any of the following occur:
1. A change of reagent lots for chemically or physically active or critical components,
unless the laboratory can demonstrate that the use of different lots does not affect
the accuracy of patient/client test results, and the range used to report patient/client
test data
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2. If QC materials reflect an unusual trend or shift or are outside of the laboratory's

acceptable limits and other means of assessing and correcting unacceptable control
values fail to identify and correct the problem
3. After major preventive maintenance or change of a critical instrument component
4. When recommended by the manufacturer
✓ Written policy defining the method, frequency and acceptability criteria for AMR verification
MIC.65170 AMR Verification Phase II

Verification of the analytical measurement range (AMR) is performed with matrix-
appropriate materials, which at a minimum, include the low, mid and high range of the
AMR, and the process is documented.
NOTE: If the materials used for calibration or for calibration verification include low, midpoint,
and high values that are near the stated AMR, and if calibration verification data are within the
laboratory's acceptance criteria, the AMR has been verified; no additional studies are required. If
the calibration and/or calibration verification materials do not span the full AMR, or the laboratory
extends the AMR beyond the manufacturer's stated range, the AMR must be verified by assaying
materials reasonably near the lowest and highest values of the AMR.
Calibration, calibration verification, and verification of the analytical measurement range (AMR)
are required to substantiate the continued accuracy of a test method. The CLIA regulations use
the term "calibration verification" to refer to both verification of correct method calibration and
verification of the analytical measurement range. This Checklist uses separate terms to identify
two distinct processes that are both required for good laboratory practice.
✓ Written procedure for AMR verification defining the types of materials used and acceptability
criteria consistent with manufacturer's instructions
REFERENCES
amendments of 1988; final rule. Fed Register. 2003(Jan 24):3707 [42CFR493.1255]
QUALITY CONTROL
Controls are samples that act as surrogates for patient/client specimens. They are processed like a patient/client
sample to monitor the ongoing performance of the entire analytic process in every run.
Qualitative molecular tests typically include positive and negative controls and, in some instances, a sensitivity
control to show that low level target is detectable. Quantitative tests typically include a negative control and
at least two levels of control at relevant decision points to verify that calibration status is maintained within
acceptable limits.
● Sampling of molecular microbiology QC policies and procedures
● Sampling of molecular microbiology QC records
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● How would you investigate results of negative controls that test as positive or
equivocal?
● What is your course of action when monthly precision data changes significantly from
the previous month's data?

determine if the steps taken follow the laboratory procedure for corrective action
● Use QC data to identify tests that utilize internal quality control processes to confirm
that any individualized quality control plan (IQCP) is used as approved by the
laboratory director
**REVISED** 08/17/2016
MIC.65200 Daily QC - Molecular-based Testing Phase II
For molecular-based quantitative and qualitative tests, controls are run at least daily, or
more frequently if specified in manufacturer's instructions, laboratory procedure, or the
CAP Checklist, and when changes occur that may impact patient results.
NOTE: The laboratory must define the number and type of quality control used and the frequency
of testing in its quality control procedures. Control testing is not required on days when patient
testing is not performed.
Controls must be run prior to resuming patient testing when changes occur that may impact
patient results, including after a change of analytically critical reagents, major preventive
maintenance, change of a critical instrument component, or with software changes, as
appropriate.
Daily quality control must be run as follows:
■ Quantitative tests - three controls at least daily, including a negative control, a low-
positive control and a high-positive control, except where a specific exception is given in
this checklist
■ Qualitative tests - a positive and negative control at least daily
Controls should verify assay performance at relevant decision points. The selection of these
points may be based on clinical or analytical criteria.
If an internal quality control process (e.g. electronic/procedural/built-in) is used instead of an
external control material to meet daily quality control requirements, the laboratory must have an
individualized quality control plan (IQCP) approved by the laboratory director to address the use
of the alternative control system. Please refer to the Individualized Quality Control Plan section of
the All Common Checklist for the eligibility of tests for IQCP and requirements for implementation
and ongoing monitoring of an IQCP.
Controls must assess adequacy of extraction and amplification, e.g. positive and negative
controls that go through the entire testing process.
● Laboratories performing tests using an IQCP approved by the laboratory director
may define their own quality control procedures to monitor the extraction and
amplification phases based on the risk assessment performed by the laboratory and the
manufacturer's instructions.
● If an IQCP is not in place that monitors the extraction and amplification processes, the
following must be followed:
1. An extraction control must be used for each run (positive controls fulfill this
requirement).
2. If the samples from an extraction batch are tested over multiple amplification runs,
each amplification run (as defined by the laboratory) must have its own amplification
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control. A single extraction control need only be tested in one of the amplification
runs.
3. If samples from multiple extraction batches are tested in a single amplification run,
each extraction batch needs an extraction control. All extraction controls must be
tested in a single amplification run. A single amplification control is sufficient.
✓ Written QC procedures
✓ Records of QC results including external and electronic/procedural/built-in control
systems AND
✓ Manufacturer's product insert or manual
REFERENCES
1) Clinical and Laboratory Standards Institute (CLSI). Statistical Quality Control for Quantitative Measurement Procedures: Principles
and Definitions; Approved Guideline. 4th ed. CLSI document C24-ED4. Clinical and Laboratory Standards Institute, Wayne, PA,
2016.
2) Ye JJ, et al. Performance evaluation and planning for patient/client-based quality control procedures. Am J Clin Pathol.
2000;113:240-248
MIC.65220 Multiplex QC Phase II

For multiplex tests, controls for each analyte are either included in each run or rotated so
that all analytes are tested periodically.
✓ Written policy defining multiplex test QC AND
✓ Records of multiplex test QC
MIC.65230 QC Acceptability Limits Phase II

Acceptability limits are defined for all control materials and standards.
NOTE: Acceptability limits must be defined for all control materials and standards. These controls
must be appropriate for the range of sensitivities tested and should, ideally, focus on result
ranges that are near clinical decision points.
✓ Written QC policy(s) defining acceptability limits
MIC.65240 QC Statistics Phase I

For quantitative assays, quality control statistics are calculated monthly to define analytic
imprecision and to monitor trends over time.
NOTE: The laboratory must use statistical methods such as calculating SD and CV monthly to
evaluate variance in numeric QC data.
✓ Written policy for monitoring of analytic imprecision including statistical analysis of data
REFERENCES
1) Mukherjee KL. Introductory mathematics for the clinical laboratory. Chicago, IL: American Society of Clinical Pathology, 1979:81-94
2) Barnett RN. Clinical laboratory statistics, 2nd ed. Boston, M; Little, Brown, 1979
3) Weisbrodt IM. Statistics for the clinical laboratory. Philadelphia, PA: JB Lippincott, 1985
4) Matthews DF, Farewell VT. Understanding and using medical statistics. NY, NY: Karger, 1988
5) Department of Health and Human Services, CMS. Clinical laboratory improvement amendments of 1988; final rule. Fed Register.
2003(Jan 24):7146 [42CFR493.1256(d)(10)(i)
6) Ross JW, Lawson NS. Analytic goals, concentrations relationships, and the state of the art of clinical laboratory precision. Arch
Pathol Lab Med. 1995;119:495-513
7) Clinical and Laboratory Standards Institute (CLSI). Statistical Quality Control for Quantitative Measurement Procedures: Principles
and Definitions; Approved Guideline. 4th ed. CLSI document C24-ED4. Clinical and Laboratory Standards Institute, Wayne, PA,
2016.
8) Brooks ZC, et al. Critical systematic error used of varied QC rules in routine chemistry. Clin Chem. 2000;46:A70
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MIC.65250 Inhibition Assessment Phase II

For assays without an internal control, the laboratory has a procedure to assess inhibition
for each specimen type.
NOTE: Documentation of an acceptable inhibition rate may be provided by the manufacturer. If

not, laboratories may test for inhibition by spiking an aliquot of the clinical specimen with target
nucleic acid. This practice can be discontinued once the laboratory accumulates sufficient data
showing that the inhibition rate falls within acceptable limits.
This requirement does not apply to probe-based solution hybridization methods (e.g. Gen-Probe
AccuProbe) performed without nucleic acid amplification.
REFERENCES
1) Ballagi-Pordany A, Belek S. Mol Cell Probes 1996 Jun 10(3):159-64
MIC.65260 Isolation/Preparation Phase II

The adequacy of nucleic acid isolation/preparation procedures is evaluated.
NOTE: Adequacy of nucleic acid isolation/preparation procedures (manual or automated) must

be evaluated with each assay by the use of positive and negative controls run in parallel with
patient samples. To the extent possible, controls must be processed through all steps of the
assay, including the extraction phase.
✓ Written procedure for evaluating adequacy of nucleic acid AND
✓ Records of controls used to assess adequacy
MIC.65270 Qualitative Cut-Off - Laboratory-Developed Test Phase II

For qualitative tests that use a cut-off value to distinguish positive from negative, the cut-
off value is established initially, and verified with every change in lot or at least every six
months.
NOTE: The limit of detection that distinguishes a positive from a negative result must be
established or verified when the test is initially placed in service, and verified with every change
in lot (e.g. new master mix), instrument maintenance, or at least every six months thereafter.
Note that a low-positive control that is close to the limit of detection can satisfy this checklist
requirement, but must be external to the kit (e.g. weak-positive patient sample or reference
material prepared in appropriate matrix).
✓ Written procedure for initial establishment and verification of the cut-off value AND
✓ Records of initial establishment and verification at defined frequency
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REAGENTS
● Sampling of reagents/controls (storage, designated pre- and post-amplification )
● How do you verify new multiplex lots/shipments?
MIC.65300 Reagent Storage Phase II

All test reagents and controls are stored properly and in a manner which minimizes target
DNA/RNA contamination and degradation.
NOTE: Pre- and post-amplification reagents and controls must be stored under appropriate
temperature and conditions in designated pre- and post-amplification areas. Temperature-
sensitive reagents and/or controls may not be stored in frost-free freezers, unless either of the
following conditions are met: 1) Reagent/control materials are kept in thermal containers and
the laboratory can demonstrate that the function of these materials is not compromised; or 2)
Freezer temperature is monitored by a continuous monitoring system, or a maximum/minimum
thermometer.
Patient samples may be stored in a frost free freezer only if the temperature is monitored in
accordance with (2), above.
✓ Written policy defining storage requirements for reagents and controls
REFERENCES
1) Borst A, Box AT, Fluit AC. False-positive results and contamination in nucleic acid amplification assays: suggestions for a prevent
and destroy strategy. Eur J Clin Microbiol Infect Dis. 2004 Apr;23(4):289-99. Epub 2004 Mar 10. Review. PMID: 15015033 [PubMed
- indexed for MEDLINE]
**REVISED** 08/17/2016
MIC.65320 New Reagent Lot - Multiplex Tests Phase II
For multiplex tests, at least two analytes are individually verified for each new shipment
and lot, and the analytes verified are periodically rotated.
NOTE: A multiplex test simultaneously detects a defined set of analytes (e.g. two or more
pathogen-specific nucleic acid sequences) from a single run or cycle of the assay. Although
a sample of analytes (at least two) may be used to verify each lot and shipment, the analytes
verified must be rotated periodically as defined in laboratory procedure to assess all analytes in
the multiplex test over time.
✓ Written procedure for new lot/shipment verification of each multiplex test AND
✓ Records of new lot and shipment verification
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**REVISED** 08/21/2017
MIC.65330 Current Primers/Probes - Laboratory-Developed Test Phase II
For tests developed by the laboratory, there are written policies and procedures to
evaluate nucleic acid tests for compatibility with currently circulating microbial strains.
NOTE: This can include, but is not limited to in silico analysis of compatibility of primers and
probes with their intended targets, surveying the literature for evidence of problems with the
assay or description of a discovered target variation that might affect test performance of the
assay. The performance of the assays in use should be assessed against newly described
variants (e.g. Influenza H1N1, EV-D68) if they occur in the patient population served by the
laboratory.
**REVISED** 08/21/2017
MIC.65340 Probe Characteristics - Laboratory-Developed Test Phase II
Information regarding the nature of any probe or primer used in a laboratory-developed
test is sufficient to permit interpretation and troubleshooting of test results.
✓ Records of probe details including oligonucleotide sequence, target, concentration, or purity,
as applicable
INSTRUMENTS
● Sampling of thermocycler well accuracy records
MIC.65400 Thermocycler Temperature Checks Phase II

Individual wells (or a representative sample thereof) of thermocyclers are checked for
temperature accuracy before being placed in service and at least annually thereafter.
NOTE: A downstream measure of well-temperature accuracy (such as productivity of

amplification) may be substituted to functionally meet this requirement. For closed systems
this function should be performed as a component of the manufacturer-provided preventive
maintenance.
✓ Written procedure for verification of thermocycler accuracy AND
✓ Records of thermocycler verification
REFERENCES
1) Saunders GC, et al. Interlaboratory study on thermal cycler performance in controlled PCR and random amplified polymorphic DNA
analyses. Clin Chem. 2001;47:47-55
2) Clinical and Laboratory Standards Institute. Establishing Molecular Testing in Clinical Laboratory Environments; Approved Guideline.
CLSI Document MM19-A. Clinical and Laboratory Standards Institute, Wayne, PA, 2011.
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PROCEDURES & TESTS
● Sampling of test policies and procedures
● Sampling of temperature incubation logs
● Sampling of molecular microbiology policies for analytic interpretation
● Physical containment practices (frequent glove change, separate manipulation of pre-

and post-specimens, dedicated pipettes)
● What is your course of action when the incubation temperature is out of range?
● What follow-up action is taken when a negative result is obtained for Group B strep
tests using direct DNA probes?
MIC.65500 Carryover Phase II

Nucleic acid amplification procedures (e.g. PCR) use appropriate physical containment
and procedural controls to minimize carryover (false positive results).
NOTE: This item is primarily directed at ensuring adequate physical separation of pre- and post-
amplification samples to avoid amplicon contamination. The extreme sensitivity of amplification
systems requires that the laboratory take special precautions. For example, pre- and post-
amplification samples should be manipulated in physically separate areas; gloves must be worn
and frequently changed during processing; dedicated pipettes (positive displacement type or with
aerosol barrier tips) must be used; and manipulations must minimize aerosolization. Enzymatic
destruction of amplification products is often helpful, as is real-time measurement of products to
avoid manual manipulation of amplification products.
✓ Written procedure that defines the use of physical containment and procedural controls as
applicable to minimizing carryover
REFERENCES
1) Kwok S, Higuchi R. Avoiding false positives with PCR. Nature 1989;339:237-238
2) Clinical and Laboratory Standards Institute. Collection, Transport, Preparations, and Storage of Specimens for Molecular Methods;
Approved Guideline. CLSI Document MM13-A. Clinical and Laboratory Standards Institute, Wayne, PA, 2005.
3) Clinical and Laboratory Standards Institute. Establishing Molecular Testing in Clinical Laboratory Environments; Approved Guideline.
CLSI Document MM19-A. Clinical and Laboratory Standards Institute, Wayne, PA, 2011.
4) Clinical and Laboratory Standards Institute. Molecular Diagnostic Methods for Infectious Diseases; Approved Guideline. 3
rd ed. CLSI
document MM03-ED3. Clinical and Laboratory Standards Institute, Wayne, PA, 2015.
MIC.65520 Temperature Range Defined Phase II

For each step of the procedure all incubation temperatures are defined and recorded.
NOTE: For some instruments this function is performed automatically by software provided by
the manufacturer.
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MIC.65530 Incubations - Manufacturer's Specifications Phase II

Incubations (reactions) performed using baths/blocks/instruments meet manufacturer's
specifications.
NOTE: Bath/blocks/instruments must be able to maintain the appropriate temperature throughout

the incubation (reaction) within the range specified by the manufacturer of the assay.
✓ Written procedure to monitor incubation performance consistent with manufacturer's
specifications
MIC.65540 Nucleic Acid Extraction/Purification Phase II

Nucleic acids are extracted and purified by validated methods.
NOTE: These can include methods reported in the literature, an established commercially
available kit or instrument, or a laboratory-developed method.
✓ Records to support nucleic acid extraction/purification is performed by a validated method
**REVISED** 08/21/2017
MIC.65550 Melting Temperature - Laboratory-Developed Test Phase II
For laboratory-developed tests that generate a result based on a melting temperature
(Tm), appropriately narrow temperature ranges (+/- 2.5ºC) are defined and recorded each
day of use.
MIC.65560 Analytic Interpretation Phase II

There are written policies for analytic interpretation of results, as applicable.
MIC.65570 Calculating Quantitative Values Phase II

For quantitative molecular tests, methods for calculating quantitative values are
adequately described and units clearly documented.
MIC.65580 Group B Screening - Non-amplified DNA Probe Phase II

Negative results obtained for Group B streptococcus intrapartum screening by a non-
amplified DNA probe are followed up with a selective broth culture method.
NOTE: Direct DNA probing is insufficiently sensitive to detect light colonization and is therefore
not adequate to replace culture based prenatal screening or to use in place of risk based
approaches when culture results are unknown at the time of labor. An adequate rapid intrapartum
test must be as sensitive as culture of vaginal and rectal swabs inoculated into selective broth
media.
✓ Written policy requiring follow-up testing for negative Group B performed by non-amplified
DNA probe
REFERENCES
1) Prevention of Perinatal Group B Streptococcal Disease: Revised Guidelines form the CDC, 2010. Morbidity and Mortality Weekly
Report http://www.cdc.gov/mmwr/preview/mmwrhtml/rr5910a1.htm
2) Prevention of Early-Onset Group B Streptococcal Disease in Newborns. 2011. The American College of Obstetricians and
Gynecologists Committee on Obstetric Practice Opinion 485. http://www.acog.org/~/media/Committee%20Opinions/Committee
%20on%20Obstetric%20Practice/co485.pdf?
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MIC.65590 Group B Screening - Amplified Method Phase II

A pre-enrichment step using a selective broth enrichment culture is performed
for antepartum (35-37 weeks gestation) vaginal/rectal swab screening for Group B
streptococci (GBS) colonization by nucleic acid amplification testing (NAAT).
NOTE: If direct sample testing is performed, without the broth enrichment step, all antepartum
samples testing negative for GBS must be followed up with a selective enrichment broth step in
conjunction with culture or NAAT testing.
The utility of NAAT assays for intrapartum testing (i.e. during active labor) remains unsettled. If
used, it is recommended that testing only be considered for women not appropriately screened
at 35-37 weeks and for whom no other clinical risk factors related to neonatal GBS infection are
present during labor.
REFERENCES
1) Prevention of Perinatal Group B Streptococcal Disease: Revised Guidelines form the CDC, 2010. Morbidity and Mortality Weekly
Report http://www.cdc.gov/mmwr/preview/mmwrhtml/rr5910a1.htm.
2) Prevention of Early-Onset Group B Streptococcal Disease in Newborns. 2011. The American College of Obstetricians and
Gynecologists Committee on Obstetric Practice Opinion 485. http://www.acog.org/~/media/Committee%20Opinions/Committee
%20on%20Obstetric%20Practice/co485.pdf?
MIC.65600 M. tuberculosis Molecular Testing Phase II

When performing molecular testing for the detection of M. tuberculosis directly from
clinical specimens, culture is performed on all samples regardless of the molecular test
result.
✓ Patient reports or worksheets
ELECTROPHORESIS
● Sampling of electrophoresis test procedures
● Sampling of melting temperature record monitoring
● Autoradiographs/gel photographs (low background, clear signal absence of bubbles)
● How is the endpoint of gel electrophoresis determined?

● What criteria are used to interpret electrophoretic gels?
MIC.65700 Molecular Weight Markers Phase II

Known molecular weight markers that span the range of expected bands are used for
each electrophoretic run.
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✓ Records of appropriate markers for each run
MIC.65720 Visual/Fluorescent Markers Phase II

Visual or fluorescent markers are used to determine the endpoint of gel electrophoresis.
MICROBIAL IN SITU HYBRIDIZATION (ISH)
● Sampling of ISH QC policies and procedures
● Sampling of ISH QC records
● What is your course of action when ISH results do not correlate with culture findings?
MIC.65800 ISH QC Phase II

Appropriate positive and negative controls are run in parallel and results recorded for
each microbial in situ hybridization (ISH) analysis.
NOTE: Laboratories should refer to the manufacturer’s guidelines for the selection of appropriate
controls. Quality control must be performed with every run, independent of the number of
samples tested (i.e. one sample or batch of several samples)
✓ Written policy for ISH QC consistent with manufacturer's guidelines
REFERENCES
1) Stefano, K., and J. J. Hyldig-Nielsen. 1997. Diagnostic applications of PNA oligomers. In S. A. Minden and L. M. Savage (ed.),
Diagnostic gene detection & quantification technologies. IBC Library Series, Southborough, Mass
2) Perry-O'Keefe, H., S. Rigby, K. Oliveira, D. Sorensen, H. Stender, J. Coull, and J. J. Hyldig-Nielsen. 2001. Identification of indicator
microorganisms using a standardized PNA FISH method. J. Microbiol. Methods 47:281-292.[CrossRef][Medline]
3) Oliveira, K., S. M. Brecher, A. Durbin, D. S. Shapiro, D. R. Schwartz, P. C. De Girolami, J. Dakos, G. W. Procop, D. Wilson, C. S.
Hanna, G. Haase, H. Peltroche-Llacsahuanga, K. C. Chapin, M. C. Musgnug, M. H. Levi, C. Shoemaker, and H. Stender. 2003.
Direct identification of Staphylococcus aureus from positive blood culture bottles. J. Clin. Microbiol. 41:889-891.[Abstract/Free Full
Text]
MIC.65820 QC Corrective Action Phase II

Corrective action is documented when microbial ISH (in situ hybridization) results do not
correlate with culture findings.
NOTE: Discordant findings should be promptly investigated for potential false positive or false
negative results from reagent failure, technical error, interpretive error or cross-reactivity of
probes.
REFERENCES
1) Stefano, K., and J. J. Hyldig-Nielsen. 1997. Diagnostic applications of PNA oligomers. In S. A. Minden and L. M.
2) Perry-O'Keefe, H., S. Rigby, K. Oliveira, D. Sorensen, H. Stender, J. Coull, and J. J. Hyldig-Nielsen. 2001. Identification of indicator
microorganisms using a standardized PNA FISH method. J. Microbiol. Methods 47:281-292.[CrossRef][Medline]
3) Oliveira, K., S. M. Brecher, A. Durbin, D. S. Shapiro, D. R. Schwartz, P. C. De Girolami, J. Dakos, G. W. Procop, D. Wilson, C. S.
Hanna, G. Haase, H. Peltroche-Llacsahuanga, K. C. Chapin, M. C. Musgnug, M. H. Levi, C. Shoemaker, and H. Stender. 2003.
95 of 97
Direct identification of Staphylococcus aureus from positive blood culture bottles. J. Clin. Microbiol. 41:889-891.[Abstract/Free Full
Text]
SEQUENCING
The requirements in this section apply to a variety of methods that can be used for sequencing (e.g. Sanger
sequencing, pyrosequencing, next generation sequencing (NGS). If NGS methods are used for infectious
disease related testing (e.g. sequences for specific organisms or taxonomic groups, assignment of drug
resistance sequences, assignment of pathogenicity markers, or assignment of host response markers), the
requirements in the Next Generation Sequencing section of the Molecular Pathology Checklist must be used in
conjunction with these requirements for inspection.
● Sampling of sequencing policies and procedures
● Manufacturer's interpretive software (most current version)
● How does your laboratory detect cross-contamination of samples/amplicons? What

action would you take if cross contamination is suspected?
MIC.65900 Sequencing Data Criteria Phase II

There are written criteria for the acceptability and interpretation of primary sequencing
data.
MIC.65920 Sequencing Data Interpretation Phase II

There is a policy to assure that appropriate databases are used and updated for the
interpretation of sequencing data.
NOTE: Data bases should be comprehensive and current.
MIC.65940 Alternative Sequencing Interpretive Databases Phase II

If the laboratory uses alternative sequence interpretive databases, either alone or in
conjunction with manufacturer’s software, the alternative databases have been validated
for the interpretation of the sequence data.
NOTE: This validation can be completed using published literature that evaluates the
interpretation of the sequence data (for example the ISA-USA resistance interpretation
guidelines). If the use of alternative data bases is done by the clinician after laboratory reporting
of sequence interpretation, this validation is not necessary.
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✓ Records of validation study if alternative interpretive databases are utilized, if applicable
MIC.65960 Sample/Amplicon Contamination Phase II

Procedures prevent or detect potential cross-contamination of samples and/or amplicons.
NOTE: Examples are the use of negative controls in each batch, the manufacturer’s use of Uracil
N-glycosylase (UNG), or the fingerprinting program provided by the manufacturer.
MIC.65980 Sample/Amplicon Contamination Phase II

If results of fingerprint analysis or negative control indicate a potential for sample and/
or amplicon contamination, the laboratory has a written procedure in place to investigate
and resolve the problem.
MIC.66000 Sequence Data Correlation Phase II

The sequence data are correlated with available phenotypic data.
✓ Records of result review including correlation with phenotypic data
RESULTS REPORTING
● Sampling of test reports (test methodology, clinical interpretation)
● Sampling of patient tests reports performed with Class I analyte specific reagents
(ASRs) including appropriate disclaimer
MIC.66100 Final Report Phase I

The final report includes a summary of the test method and information regarding clinical
interpretation if appropriate.
NOTE: For example, HIV-1 viral load results may vary significantly depending upon the test
method used; including the test method in the report is important information for interpreting the
results.
**REVISED** 08/21/2017
MIC.66120 ASR Disclaimer Phase II
If patient testing is performed using Class I analyte-specific reagents (ASRs) obtained or
purchased from an outside vendor, the patient report includes the disclaimer statement
required by federal regulations.
NOTE: ASRs are antibodies, both polyclonal and monoclonal, specific receptor proteins, ligands,
nucleic acid sequences, and similar reagents which, through specific binding or chemical reaction
with substances in a specimen, are intended for use in a diagnostic application for identification
and quantification of an individual chemical substance or ligand in biological specimens.
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An ASR is the active ingredient of a laboratory-developed test system. Class I ASRs are not
subject to preclearance by the US Food and Drug Administration (FDA) or to special controls by
the FDA.
If the laboratory performs patient testing using Class I ASRs, federal regulations require that the
following disclaimer accompany the test result on the patient report: "This test was developed
and its performance characteristics determined by (laboratory name). It has not been cleared or
approved by the US Food and Drug Administration."
The CAP recommends additional language, such as "The FDA does not require this test to go
through premarket FDA review. This test is used for clinical purposes. It should not be regarded
as investigational or for research. This laboratory is certified under the Clinical Laboratory
Improvement Amendments (CLIA) as qualified to perform high complexity clinical laboratory
testing."
The disclaimer is not required for tests using reagents that are sold in kit form with other
materials and/or an instrument, and/or with instructions for use, and/or when labeled by the
manufacturer as Class I for in vitro diagnostic use (IVD), Class II IVD, or Class III IVD.
The laboratory must establish the performance characteristics of tests using Class I ASRs in
accordance with the Method Performance Specifications section of the All Common Checklist.
The laboratory may put a single ASR disclaimer on the patient report for all microbiology studies
collectively used in a particular case. Separately tracking each reagent used for a case and
selectively applying the disclaimer to only the class I ASRs is unnecessary.
REFERENCES
1) Department of Health and Human Services, Food and Drug Administration. Medical devices; classification/reclassification; restricted
devices; analyte specific reagents. Final rule. Fed Register. 1997(Nov 21);62243-45 [21CFR809, 21CFR864]
2) Caldwell CW. Analyte-specific reagents in the flow cytometry laboratory. Arch Pathol Lab Med. 1998;122:861-864
3) Graziano. Disclaimer now needed for analyte-specific reagents. Northfield, IL: College of American Pathologists CAP Today.
1998;12(11):5-11
LABORATORY SAFETY
The inspector should review relevant requirements from the Safety section of the Laboratory General checklist,
to assure that the molecular testing section is in compliance. Please elaborate upon the location and the details
of each deficiency in the Inspector's Summation Report.
The following requirement pertains specifically to the molecular microbiology section.
● Sampling of molecular microbiology specimen handling and processing policies

There are written policies for the safe handling and processing of samples from patients
with suspected infections due to avian influenza, SARS, or similar emerging pathogens.

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ON-LINE CHECKLIST AVAILABILITY

SUMMARY OF CHECKLIST EDITION CHANGES

NEW Checklist Requirements

Requirement Effective Date

REVISED Checklist Requirements

Requirement Effective Date

DELETED/MOVED/MERGED Checklist Requirements

Requirement Effective Date

MIC.00350 PT Extent of Testing Phase II

QUALITY MANAGEMENT AND QUALITY CONTROL

● Select several occurrences in which QC is out of range and follow records to

MIC.10060 QC - Waived Tests Phase II

NOTE: Quality control must be performed according to manufacturer's instructions. Testing

MIC.10070 QC Corrective Action - Waived Tests Phase II

MIC.11015 QC Handling Phase II

MIC.11016 Commercial Product - QC Phase II

MIC.11017 QC Confirmation of Acceptability Phase II

MIC.11018 QC Corrective Action Phase II

MIC.11020 Monthly QC Review Phase II

NOTE: "Performance" includes elements of accuracy, precision, and clinical discriminating

MIC.11060 Culture Result Reporting Phase I

MIC.11350 Morphologic Observation Assessment Phase II

NOTE: Suggested methods to accomplish this include:

1. Circulation of organisms with defined staining characteristics, and/or

MIC.11375 Taxonomy Changes Phase I

SPECIMEN COLLECTION AND HANDLING

● Sampling of microbiology specimens (transport media, timely delivery, labeling)

● What is your course of action when you receive unacceptable microbiology

MIC.13175 Viral Culture Specimens Phase I

MIC.13200 Requisitions Phase I

MIC.13250 Specimen Collection/Handling Phase II

NOTE: Manufacturer's recommendations must be followed when there is a delay in delivery or

MIC.13275 Specimens for Molecular Amplification Phase II

MIC.14583 Direct Antigen Test QC Phase II

MIC.15000 Preliminary Reports Phase I

MIC.15020 Azoospermic Specimen Result Reporting Phase I

INSTRUMENTS AND EQUIPMENT

● Incubators (adequate space, maintained)

MIC.16150 Pipettors and Diluters Phase II

MIC.16550 Adequate Incubators Phase I

MATRIX-ASSISTED LASER DESORPTION IONIZATION

MIC.16575 Instrument Operation Phase II

MIC.16595 Mass Spectrometer Calibration Phase II

MIC.16615 Mass Spectrometer Reagent Grade Phase II

MIC.16625 Mass Spectrometer Consumables Phase II

NOTE: For FDA-approved platforms, consumables utilized may be specified by the

MIC.18968 Agents of Bioterrorism Phase II

NOTE: Microorganisms likely to be utilized as biological weapons include Bacillus anthracis

MIC.18985 Spill Handling Phase II

MIC.19010 Bench Top Decontamination Phase II

MIC.19035 Safe Specimen Processing Phase II

MIC.19060 Biosafety Levels Phase II

MIC.19160 Biosafety Levels Phase II