STANDARD OPERATING PROCEDURE (SOP)

Legacy Pathology Laboratory, Kigali Rwanda

LEGACY COBAS E411 USE SOP

CLINICS PATHOLOGY Effective date: Pages:13
LABORATORY LEG/PATHLAB/BIO-SOP-25 --
VERS 001

Review date:

Prepared by: Verified by: Authorised by 

Names: Names :………… Name:

Title: …………………
Title::………………. Laboratory Director
Signature :……………
Signature ………………….

Date :………………… Date:…………….. Date:……………

REVISION HISTORY
Revised by Version Review date Effective date Description of change

NA 001 NA

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 STANDARD OPERATING PROCEDURE (SOP)

Legacy Pathology Laboratory, Kigali Rwanda

1. INTRODUCTION
The Roche Diagnostic Cobas e 411immunoassay system is a fully automated, software-
controlled system, multichannel analyzer for immunoassay analysis. It is designed for
both quantitative and qualitative in vitro determination of analytes by use
Electrochemiluminescence (ECL) technology.
2. DEFINITIONS AND ABBREVIATIONS
2.1 Definitions
Immunoserology: Branch of serology that study immune response.
Antibody: blood protein produced in response to and counteracting a specific antigen.
Antigen: a foreign substance, which induces immune response in the body, especially the
production of specific antibody.
ECL: is a process in which highly reactive species are generated from stable precursors at
the surface of an electrode. These highly reactive species react with one another,
producing light.
2.2 Abbreviations
SOP: Standard Operating Procedure
IMS: Immunoserology
ISE: Ion Selective Electrode
TPA: tripropylamine
Ig: Immunoglobulin
Ab: Antibody
Ag: Antigen
ECL: Electrochemiluminescence technology
FSH: Follicle stimulating hormone
LH: Luteinizing hormone
E2: Estradiol
HCG: Human Chorionic gonadotropin
AFP: alpha fetoprotein
CEA: Carcino embryogenic antigen

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CA: Cancer Antigen

PRL: Prolactin
TSH: Thyroid stimulating hormone
T3: Tri iodothyronine
T4: Tetra iodothyronine
CMV: Cytomegalovirus
HBSAG: Hepatitis B surface antigen
HCV-Ab: Hepatitis C Virus Antibody
HBEAG: Hepatitis B Enveloppe antigen
HBCAB: Hepatitis B Core antibody
B12: vitamin B12 (cyanocobalamin)
3 SCOPE
This SOP applies to all staff using Cobase411 instrument in the Qualitative and quantitative
determination of HBSAG, HBeAg, HBe-Ab, HBC-Ab, HBS-Ab,HCV-Ab, HIV Combi, Toxo
IgG, IgM, Rubella IgG and IgM, Cytomegalovirus IgG and IgM, TSH, Free T4, Free T3, T3, T4,
FSH, LH, Progesterone, Estradiol, Prolactin, Testosterone. Beta HCG total, Free HCG, Cortisol,
Total PSA, Free PSA, CA125, CA 19-9, CA15-3, CEA, AFP B12, Folic Acid and Ferritin in
plasma or serum
4 ROLES AND RESPONSIBILITIES
Person responsible Tasks
Technologist/scientist Perform tests as per procedure

Head of unit Ensure tests have been performed as per procedure

Verify and validate results before release

Quality manager Ensure that this procedure remain updated and relevant
Lab Director Overall responsibility to ensure implementation of this
procedure

5 PURPOSE OF THE EXAMINATION

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 STANDARD OPERATING PROCEDURE (SOP)

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This SOP provides the instructions for the use of Cobase411 in qualitative and quantitative
determination for Immuno-serology analytes in human Serum or Plasma to roll out infections,
fertile disorder, thyroid disorders, iron studies and tumour markers

6 PRINCIPLE AND METHOD OF THE PROCEDURE

The Cobas e411 has competitive principle for extremely small analytes, sandwich principle for
larger analytes and a bridging principle to detect antibodies in the sample.
The cobas e411 utilizes electrochemiluminescence (ECL) method. ECL is a process in which
highly reactive species are generated from the stable precursors at the surface of an electrode.
These highly reactive species react with one another and produce light.
The development of ECL Immunoassays is based on the use of a ruthenium (II)-tris(bipyridyl)
complex and tripropylamine (TPA). The final chemiluminescent product is formed during the
detection step.
The chemiluminescent reactions that lead to the emission of light from the ruthenium complex
are initiated electrically by applying voltage to the immunological complexes that are attached to
the streptavidin-coated microparticles.
7 PERFORMANCE CHARACTERISTICS
First class performance: The sensitivity, reliability and reproducibility of results is
traceable to ECL technology. More than 100 assays available, up to 18 assays can be run at a
throughput of up to 86 tests /hr. 9 min STAT applications for emergency sample assays.
Intelligent sample workflow: Carryover-free pipetting of samples to avoid cross-
contamination; automated sample rerun capability; continuous random access and flexible
STAT priority settings.
Unique reagent concept: Convenient and error-free handling of cobas e packs; economic
usage with high stabilities and convenient kit sizes.
High system reliability: More than 25,000 analyzers installed worldwide, high uptime
>99%; low system maintenance time.
8 TYPE OF SAMPLE
Serum
Plasma

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 STANDARD OPERATING PROCEDURE (SOP)

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9 PATIENT PREPARATION
This is done as per procedure on sample collection, transportation, reception and storage

10 TYPE OF CONTAINER AND ADDITIVES

Red top tubes
EDTA tubes
11 REQUIRED EQUIPMENT AND REAGENTS
Equipment & supplies
Cobase411 and its accessories
Centrifuge
Micropipettes
Refrigerator (2 – 8oC)
Sample cups
Assay cups
Assay tips
ISE cleaning solution,
Reagents
Testing reagents: ready to use reagent packs labelled with a specific test name
Calibrators: Each reagent pack has it own calset.
Controls: Precicontrol Universal for wide range of analytes, precicontrol tumor marker,
other specific control material for infectious disease tests.
Consumables: Syswash, Procell, clean cell, Deionised water

12 ENVIRONMENTAL AND SAFETY CONTROLS

All specimens are handled using appropriate safety procedures as identified in the
Laboratory Safety Manual.
Room temperature for CCH laboratory shall be 20 to 25 degrees. The temperature shall be
checked twice a day (morning and evening) and recorded in the temperature-monitoring log.
The CCH benches are disinfected every morning using 5% hydrochloride solution.
Windows in CCH shall be closed at all times.
Waste disposal is done as per waste management guideline

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 STANDARD OPERATING PROCEDURE (SOP)

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13. CALIBRATION PROCEDURES

 The Cobase411analyzer shall be calibrated and measurements traceable to SI units
 Calibrator positions are selected manually in the analyzer.
 Calibration is done at every change in reagent lot number and as required following
quality control procedures.

Note: Calibrators take the same names as their reagents + level1 & level2

 Put the calibrator material at room temperature 15- 30 minutes

 On the screen, select the calibration menu, Calibrator sub menu
 Select the test level 1 and assign the position
 Select the test level 2 and assign the position
 Lift up the probe guard
 Put the calibrator materials in their specific positions on sample disk
 Lift down the probe guard
 Press START button
 Calibration results are printed automatically
 Check if calibration was generated
 Validate the results and keep the document

 For Hormones, tumor markers and Vitamins

Note: Each calibrator takes the name of reagent it calibrates + Calset word( e.g:
Testosterone Calset)
 Put the calibrator material at room temperature 15- 30 minutes
 Prepare the reagent respecting the volumes of distilled water mentioned on the container
 leave it for 30 minutes for at room temperature
 Transfer the working reagent into the proper cups
 On the screen, select the calibration menu, Install sub menu

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 Put the Card containing the calibration informations into the scanner position
 Click on Scan BC Card button, the analyzer scan automatically the card and goes in
standby mode.
 Calibrator sub menu
 Select the test level 1 and assign the position
 Select the test level 2 and assign the position
 Lift up the probeguard
 Put the calibrator materials in their specific positions on sample disk
 Lift down the probe guard
 Press START button
 Calibration results are printed automatically
 Check if calibration was generated
 Validate the results and keep them in specified file
13 PROCEDURAL STEPS
14.1. Special precautions

 The users should always ensure that the external water supply level is
enough
 The users should all the time check for error messages and printed them
out.
 The users must be able to identify the colour due to the message
displayed ( Reference to e411 Operator’s manual,Chapter 5, B-12.
 The users should all the time check for level of waste reservoir and waste
box, discard waste when full.
 The reagents and consumables should be kept in appropriate condition.
 The users should always communicate immediately the critical results to
the clinicians

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14.2 Starting up the Cobas e 411 instrument

 Check if the machine is connected to the electricity supply
 Open the cover, verify the following:
 The volume of syswash and deionized water,
 pro-cell and clean-cell reagents,
 Sample tips and sample cups,
 liquid waste and solid waste container,
 The probe guard must be lifted down and sipper nozzle closed.
 Lift up the circuit breaker to start-up the machine.
 Switch on the operation switch.
 Switch on the monitor
 Wait until the machine displays the window, …. user name and password
 Fill in the username and password
 Wait until the machine goes in the stand-by mode.
14.3 Loading reagents
 Remove the reagents from the refrigerator according to the requested tests.
 Wait 15 minutes for the reagents to take the room temperature.
 Open the reagents disk cover and put the reagents opened, facing to the right side
randomly and recover the reagents disk.
 Go to the reagents menu click to the reagents scan, after scanning reagents the machine
will return to the standby mode.

14.4 Reagent calibration

Reagents are calibrated as per calibration procedures (point 13 Above.)
14.5 Running Samples
 Centrifuge the samples
 Check the test requested in Open clinic system and record on bench sheet
 Label samples
 Label the sample cups
 put the serum/plasma in the labeled sample cups according to the disk positions

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 STANDARD OPERATING PROCEDURE (SOP)

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 Put them from one to the sample disk according to their numbers. (1-1; 2-2 ;…)
 After loading the samples, put at the last number the sample stop barcode the bars facing
the scanner.
 Lift down the probe guard.
 Go to the screen, select Workplace menu, test selection sub menu, routine- serum or
plasma,
 Sequence number is recorded automatically,
 Go to sample position and enter the number corresponding the patient sample and the
request and press Enter button
 Enter the identification of the patient (names) and press Enter button
 Select tests and save.
 Click start button, and the big start button appears, click it and wait the machine to
analyze all the samples,
 Results are recorded are printed/recorded on the bench sheet by patient.
 Results are verified and sent to clinicians via open clinic system.

14.6 Shutting down the e411

 Click on log off button
 Click shut down, and the machine does it itself.
 Switch off the operation switch and if you need the reagent to remain on board, don’t
switch off the circuit breaker.
 Remove the reagents and keep them in a refrigerator
 Switch off the circuit breaker.

15. QUALITY CONTROL PROCEDURES.

15.1 Internal quality control
Commercial QCs materials are used for the internal quality controls for all reagents
IQCs are done daily for the following tests: HBSAG, HCV-Ab, Tumor markers, vitamins and
hormones in general.
IQCs are done monthly for: other infectious diseases tests.

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 STANDARD OPERATING PROCEDURE (SOP)

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Failed QCs are repeated after calibration.

Control results are signed by staff on bench and reviewed by the head of unit.
Control results are filed in their specific file.

Running IQCs:
Note: some control materials used on the cobas e411 are ready for use; others need to be
reconstituted when they come lyophilized (follow the instructions on the control container).
 Put the control materials at room temperature 15- 30 minutes
 Scan the Cards containing the information of Negative and Positive controls, Normal
and Pathologic controls.
 Reconstitute the controls that came in powder and wait 30 minutes at room
temperature
 Put the controls into their containers (sample cups)
 Assign the positions for the control materials
 Put the control materials into positions on sample disk
 Press START button and wait the results to be printed automatically
 Validate the results and document them.
16. INTERFERENCES AND CROSS REACTIONS
NA
15. RESULTS CALCULATION AND MEASUREMENT OF UNCERTAINTY
The measurement of uncertainty is calculated using data from internal quality control.
The analyser automatically calculates the analyte concentration from each sample in specific unit
of measurement based on calibration data and observed data in-patient Sample.
17. BIOLOGICAL REFERENCE INTERVALS OR CLINICAL DECISION VALUES
Each and every test done on Cobas e411 has the reference value (refer to the immunoassay
test inserts and result reports).
18. REPORTABLE INTERVAL OF EXAMINATION RESULTS
Measuring ranges for each test are followed and respect as per test insert.
19. DETERMINATION OF QUANTITATIVE RESULTS OUT OF MEASUREMENT
INTERVAL

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Samples having higher concentrations are diluted considering the specific dilution mentioned by
manufacturer in the test insert. Results from samples diluted by the rerun function are
automatically multiplied by a dilution factor. (Reference to the test insert).
The lower detection limit represents the lowest measurable analyte level that can be
distinguished from zero. It is calculated as the value lying three standard deviations above that of
the lowest standard.

20. CRITICAL VALUES, WHERE APPROPRIATE

- Positive result for Qualitative determination is reported as soon as possible to the clinic.
21. LABORATORY CLINICAL INTERPRETATION
- Quantitative results are interpreted as: normal, low, high, below detectable limit, based on
reference values.
- Qualitative results are interpreted as negative or positive.
- The results should always be assessed in conjunction with the patient’s medical history,
clinical examination and other findings.
22. Potential sources of variation
- Cortisol has periodic changes as it decreases by time; sample should be taken morning
and evening.
- Early testing of infectious diseases parameters ( window period of each virus , bacteria or
parasites).
23. REFERENCES
- Cobase411 user’s operator manual
- Immunoserology test inserts

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Legacy Pathology Laboratory, Kigali Rwanda

24.SIGNATURE PAGE
We, staff members of Legacy Pathology Laboratory do hereby certify that we have read,
discussed (where applicable) and understood the content of this SOP.
We commit ourselves to abide by its spirit and shall strive to comply and make it complied with.
N° Staff Names Staff Signature Date
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17

I, ……………………………….........…..…… the Laboratory Director indicated above, do

hereby certify that all the staff, as listed above, have read, discussed and understood the SOP as
indicated herein.

Name and signature : Date:

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24. AMENDMENT PAGE

Date Amendment Amendment Page Section(s) Author

number involved

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