CRITERION™ CLED AGAR

Cat. no. C5400 CRITERION™ CLED Agar 72gm
Cat. no. C5401 CRITERION™ CLED Agar 500gm
Cat. no. C5402 CRITERION™ CLED Agar 2kg
Cat. no. C5403 CRITERION™ CLED Agar 10kg
Cat. no. C5404 CRITERION™ CLED Agar 50kg

INTENDED USE

Hardy Diagnostics CRITERION™ CLED Agar is recommended for the isolation and enumeration of microorganisms from urine.

This dehydrated culture medium is a raw material intended to be used in the making of prepared media products, which will require further processing, additional ingredients, or supplements.

SUMMARY

In the mid 1960s, Sandys and Mackey reported on the laboratory diagnosis of urinary tract infections using a new medium Sandys had developed to prevent the swarming of Proteus spp.(7,8,10) Previous culture methods used to inhibit the swarming of Proteus included adding chloral hydrate, alcohol, sodium azide, surface-active agents, boric acid, and sulfonamides to the medium.(10) However, Mackey and Sandys' modified medium replaced mannitol with lactose, discontinued the use of sucrose, increased the indicator strength of bromothymol blue and the concentration of agar, and incorporated the use of cystine in order to enhance the growth of cystine-dependent "dwarf colony" coliforms.(8) They named their final medium Cystine Lactose Electrolyte-Deficient (CLED) Agar and reported it as ideal for dip-inoculum techniques and for general urinary bacteriology and colony differentiation. CLED Agar also lacks sodium chloride, which helps in preventing the swarming of Proteus spp.

CLED Agar supports the growth of all potential urinary pathogens, and a number of contaminants such as diphtheroids, lactobacilli, and micrococci. Urine samples containing mixed flora are typical of urethral or vaginal contamination.

Research demonstrates that the best results are obtained when inoculation occurs as soon after sample collection as possible.(2,8) Otherwise, confluent or semiconfluent growth may occur when CFU counts exceed 105 per ml of urine. Historically, reliable quantitative urine cultures have been obtained using the dip-inoculum method, even when there was a time delay of several hours between inoculation of urine onto the medium and incubation.(2) Therefore, the inoculated medium may be held for 48 hours or longer, refrigerated or at room temperature, until received by a testing facility. This makes CLED Agar extremely versatile for physicians' offices, small clinical labs, or hospital wards, and eliminates the need for transport and refrigeration of patient urine specimens.

Hardy Diagnostics CRITERION™ CLED Agar is recommended for use in the spread plate technique or the dip-inoculum method for detection of bacteria in urine. CRITERION™ CLED Agar contains enzymatic digest of casein, enzymatic digest of gelatin, and beef extract, which provide nitrogen, vitamins, and carbon to support microbial growth. Lactose is added as the carbohydrate source. L-cystine is a growth supplement for cystine-dependent coliforms. Organisms capable of fermenting lactose will lower the pH and change the color of the medium to yellow. Consequently, bromothymol blue is the pH indicator. Agar acts as the solidifying agent.

FORMULA*

Gram weight per liter: 36.0gm/L
Lactose 10.0gm
Pancreatic Digest of Gelatin 4.0gm
Pancreatic Digest of Casein 4.0gm
Beef Extract 3.0gm
L-Cystine 0.128gm
Bromothymol Blue 0.02gm
Agar 15.0gm

Final pH 7.3 +/- 0.2 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.

STORAGE AND SHELF LIFE

Store the sealed bottle(s) containing dehydrated culture medium at 2-30ºC. Dehydrated culture medium is very hygroscopic. Keep lid tightly sealed. Protect dehydrated culture media from moisture and light. The dehydrated culture media should be discarded if it is not free-flowing or if the color has changed from its original light bluish-tan.

Store the prepared culture media at 2-8ºC.

PRECAUTIONS

METHOD OF PREPARATION FOR DEHYDRATED CULTURE MEDIA

1. Suspend 36gm of the dehydrated culture media in 1 liter of distilled or deionized water. Stir to mix thoroughly.

2. Heat to boiling for one minute to dissolve completely.

3. Check the pH and adjust if necessary.

4. Sterilize in the autoclave at 121ºC. for 15 minutes.

5. Cool to 45-50ºC.

6. Aseptically pour into desired sterile containers.

PROCEDURE AND INTERPRETATION OF RESULTS

For information on procedures and interpretation of results, consult listed reference or refer to the prepared media Instructions for Use (IFU) for Cat. No. G223.

LIMITATIONS

Limiting factors in low urine counts from infected patients include: rapid rate of urine flow, prior initiation of antimicrobial treatment, urine with a pH less than 5 and a specific gravity less than 1.003.(8)

The nutritional requirements of organisms vary and some strains may grow poorly or fail to grow entirely on this medium.

CRITERION™ CLED Agar is a non-selective medium. However, the growth of some Shigella species may be inhibited due to electrolyte exclusion in the formula.(6)

MATERIALS REQUIRED BUT NOT PROVIDED

Standard microbiological supplies and equipment such as autoclaves, incinerators, and incubators, etc., are not provided.

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Enterococcus faecalis
ATCC ® 29212
A 24hr 35°C Aerobic Growth; small yellow colonies
Escherichia coli
ATCC ® 25922
A 24hr 35°C Aerobic Growth; yellow colonies, opaque, center slighlty deeper yellow
Proteus mirabilis
ATCC ® 12453
A 24hr 35°C Aerobic Growth; translucent blue colonies
Staphylococcus aureus
ATCC ® 25923
A 24hr 35°C Aerobic Growth; deep yellow colonies
Pseudomonas aeruginosa
ATCC ® 27853
A 24hr 35°C Aerobic Growth; green colonies with matte surface and rough periphery
Klebsiella pneumoniae
ATCC ® 13883
A 24hr 35°C Aerobic Growth; yellow to whitish-blue colonies, mucoid

User Quality Control

Physical Appearance

CRITERION™ CLED Agar powder should appear homogeneous, free-flowing, and light bluish-tan in color. The prepared media should appear slightly opalescent, and light blue-green in color.

REFERENCES

1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Benner, E.J. 1970. Simple Disposable Method for Quantitative Cultures of Urine. Appl. Micro. Vol. 19, No. 3.

3. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

4. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

5. Koneman, E.W., et al. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott Company, Philadelphia, PA.

6. MacFaddin, J.F. 1985. Media for Isolation, Cultivation, Identification, Maintenance of Bacteria, Vol. I. Williams & Wilkins, Baltimore, MD.

7. Mackey, J.P and G.H. Sandys. 1965. Laboratory Diagnosis of Infections of the Urinary Tract in General Practice by Means of a Dip-inoculum Transport Medium. Brit. Med. J.; 2:1286-1288.

8. Mackey, J.P. and G.H. Sandys. 1966. Diagnosis of Urinary Infections. Brit. Med. J.; 1:1173.

9. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

10. Sandys, G.H. 1960. A new method of preventing swarming of Proteus spp. with a description of a new medium suitable for use in routing laboratory practice. J. Med. Lab. Technol.; 17:224.


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051616gr