M352

R
Technical Data
C.L.E.D. Agar w/ Andrade Indicator M352
Intended Use:
Recommended for isolation and differentiation of urinary pathogens on the basis of lactose fermentation.
Composition**
g/L
Ingredients
Peptone 4.000
HM peptone B # 3.000
Tryptone 4.000
Lactose 10.000
L-Cystine 0.128
Bromothymol blue 0.020
Andrade indicator 0.100
Agar 15.000
Final pH ( at 25°C) 7.5±0.2
**Formula adjusted, standardized to suit performance parameters

# - Equivalent to Beef extract
Directions
Suspend 36.25 grams in 1000 ml purified/distilled water. Heat to boiling to dissolve the medium completely. Sterilize by
autoclaving at 15 lbs pressure (121°C) for 15 minutes. Cool to 45-50°C. Mix well and pour into sterile Petri plates.
Principle And Interpretation
Sandys reported a new technique where the swarming of Proteus on an agar medium could be prevented by restricting the
electrolyte content in the culture medium (1). Sandys Medium was modified by Mackey and Sandys (2), by replacing
mannitol with lactose and sucrose and elevating the concentration of agar and bromothymol blue. The same authors further
modified this medium by retaining the lactose (deleting sucrose) and by including L-cystine for promoting the growth of
cystine-dependent dwarf coliform colony (3). This later modified medium was designated as C.L.E.D. (Cystine- Lactose-
Electrolyte-Deficient) Medium. This medium is recommended for use in urinary bacteriology, promoting the growth of all
urinary pathogens. C.L.E.D. Medium is also recommended for dip stick procedures and as dip inoculum transport medium for
urine specimens (2,3,4).
C.L.E.D. Medium was further modified by Bevis (5) by incorporation of Andrades indicator. This medium provides sharper
differentiation between lactose-fermenters (LF) and lactose-non-fermenters (NLF) (5). Addition of Andrades indicator
enhances the appearance of colony and aids in the identification of microorganisms.
At different pH values, the colour of the medium varies from the standard medium, which is well documented by Bevis (5).
pH Colour of C.L.E.D. medium
7.4 deep blue
7.0 bluish grey
6.8 pale grey
6.6 pinkish grey
6.4 bright red with whitish tinge
6.0 bright red
For better results, the medium should not be incubated for more than 24 hours because if lactose fermenters predominate, the
entire medium may turn pink masking the presence of non-lactose fermenters. Inoculate the medium immediately after urine
collection. Shigella species may not grow on this medium. Prior initiation of antibiotic therapy, low urine pH (less than 5)
etc. may result in low urine count from infected patients.
Type of specimen
Clinical:Urine sample
Please refer disclaimer Overleaf.

HiMedia Laboratories Technical Data
Specimen Collection and Handling

For clinical samples follow appropriate techniques for handling specimens as per established guidelines (6,7).
After use, contaminated materials must be sterilized by autoclaving before discarding.
Warning and Precautions
In Vitro diagnostic use only. For professional use only. Read the label before opening the container. Wear
protective gloves/protective clothing/eye protection/face protection. Follow good microbiological lab practices while
handling specimens and culture. Standard precautions as per established guidelines should be followed
while handling clinical specimens. Safety guidelines may be referred in individual safety data sheets.
Limitations
1. This medium is recommended for urine infection. Low urine count may be a result of antibiotic therapy, low pH of
urine.
2. Recovery depends on the urine count.
3. Inoculate the medium immediately after urine collection.
4. Shigella species may not grow on this medium.
5. For better results, the medium should not be incubated for more than 24 hours because if lactose fermenters
predominate, the entire medium may turn pink masking the presence of non-lactose fermenters.
Performance and Evaluation
Performance of the medium is expected when used as per the direction on the label within the expiry period when stored
at recommended temperature.
Quality Control
Appearance
Light yellow to grayish yellow homogeneous free flowing powder
Gelling
Firm, comparable with 1.5% Agar gel
Colour and Clarity of prepared medium
Greenish blue clear to slightly opalescent gel forms in Petri plates
Reaction
Reaction of 3.62% w/v aqueous solution at 25°C. pH : 7.5±0.2
pH
7.30-7.70
Cultural Response
Cultural characteristics observed after an incubation at 35-37°C for 18-24 hours
Organism Inoculum Growth Recovery Colour of
(CFU) colony
# Klebsiella aerogenes 50-100 good-luxuriant >=70% greyish green

ATCC 13048 (00175*)
Escherichia coli ATCC 50-100 good-luxuriant >=70% bright pink
25922 (00013*) with pink halo
Enterococcus faecalis 50-100 good-luxuriant >=70% orange-yellow
ATCC 29212 (00087*) or greenish
Proteus mirabilis 50-100 good-luxuriant >=70% blue-green
ATCC 25933
Staphylococcus aureus >=70% golden-yellow
50-100 good-luxuriant
subsp. aureus ATCC
25923 (00034*)
Streptococcus pyogenes 50-100 good-luxuriant >=70% greyish green
ATCC 19615
Key : *Corresponding WDCM numbers.
#- Formerly known as Enterobacter aerogenes
Storage and Shelf Life

Store between 10-30°C in a tightly closed container and the prepared medium at 20-30°C. Use before expiry date on the
label. On opening, product should be properly stored dry, after tightly capping the bottle in order to prevent lump formation
due to the hygroscopic nature of the product. Improper storage of the product may lead to lump formation. Store in dry
ventilated area protected from extremes of temperature and sources of ignition Seal the container tightly after use. Product
performance is best if used within stated expiry period.
Please refer disclaimer Overleaf.
HiMedia Laboratories Technical Data
Disposal
User must ensure safe disposal by autoclaving and/or incineration of used or unusable preparations of this product. Follow
established laboratory procedures in disposing of infectious materials and material that comes into contact with clinical
sample must be decontaminated and disposed of in accordance with current laboratory techniques (6,7).
Reference
1. Sandys, 1960, J. Med. Lab. Technol., 17:224.
2. Mackey and Sandys, 1965, Br. Med. J., 2:1286.
3. Mackey and Sandys, 1966, Br. Med. J., 1:1173.
4. Dixson J. M. S. and Clark M. A., 1968, Conc. Med. Assoc. J., 99 (15)
5. Bevis T. D., 1968, J. Med. Lab. Technol., 25:38.
6. Isenberg, H.D. Clinical Microbiology Procedures Handbook. 2nd Edition.
7. Jorgensen, J.H., Pfaller, M.A., Carroll, K.C., Funke, G., Landry, M.L., Richter, S.S and Warnock., D.W. (2015) Manual of
Clinical Microbiology, 11th Edition. Vol. 1.
Revision : 06/ 2024
HiMedia Laboratories Pvt. Limited, IVD In vitro diagnostic 30°C Storage temperature
Plot No.C-40, Road No.21Y, medical device
MIDC, Wagle Industrial Area,
Thane (W) -400604, MS, India 10°C
EC REP CEpartner4U, Esdoornlaan 13, Do not use if

CE Marking
3951DB Maarn, NL package is damaged
www.cepartner4u.eu
Disclaimer :
User must ensure suitability of the product(s) in their application prior to use. Products conform solely to the information contained in this and
other related HiMedia™ publications. The information contained in this publication is based on our research and development work and is to the best
of our knowledge true and accurate. HiMedia™ Laboratories Pvt Ltd reserves the right to make changes to specifications and information related
to the products at any time. Products are not intended for human or animal or therapeutic use but for laboratory,diagnostic, research or further
manufacturing use only, unless otherwise specified. Statements contained herein should not be considered as a warranty of any kind, expressed or
implied, and no liability is accepted for infringement of any patents.
HiMedia Laboratories Pvt. Ltd. Corporate Office : Plot No.C-40, Road No.21Y, MIDC, Wagle Industrial Area, Thane (W) - 400604, India.
Customer care No.: 022-6147 1919 Email: techhelp@himedialabs.com Website: www.himedialabs.com

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**Formula adjusted, standardized to suit performance parameters

Please refer disclaimer Overleaf.

Specimen Collection and Handling

# Klebsiella aerogenes 50-100 good-luxuriant >=70% greyish green

Storage and Shelf Life

Revision : 06/ 2024

EC REP CEpartner4U, Esdoornlaan 13, Do not use if

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