CRITERION™ LOWENSTEIN MEDIUM BASE

Cat. no. C7380 CRITERION™ Lowenstein Medium Base 74.4gm
Cat. no. C7381 CRITERION™ Lowenstein Medium Base 500gm
Cat. no. C7382 CRITERION™ Lowenstein Medium Base 2kg
Cat. no. C7383 CRITERION™ Lowenstein Medium Base 10kg
Cat. no. C7384 CRITERION™ Lowenstein Medium Base 50kg

INTENDED USE

Hardy Diagnostics CRITERION™ Lowenstein Medium Base, when supplemented with egg and glycerol, is used for the cultivation, isolation, and differentiation of Mycobacterium spp.

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

The original formulation of Lowenstein Jensen media was developed by Lowenstein who incorporated congo red and malachite green to inhibit unwanted bacteria.(6,7) The present formulation is based upon Jensen's modification. Jensen's version eliminates congo red and uses a moderate concentration of malachite green to prevent growth of the majority of contaminants surviving decontamination of the specimen. This formulation also encourages the earliest possible growth of mycobacteria.

Fresh eggs are added aseptically to this media. During heating, the egg albumin coagulates, thus providing a solid surface for inoculation. Nitrogen, fatty acids, and proteins are supplied by egg and asparagine. Glycerol serves as a carbon source and is favorable to the growth of the human type tubercle bacillus while being unfavorable to the bovine type. Malachite green acts as an inhibitory agent toward microorganisms other than mycobacteria.(8)

FORMULA

Gram weight per 600ml: 37.2gm/600ml
Potato Starch 30.0gm
Asparagine 3.6gm
Monopotassium Phosphate 2.4gm
Magnesium Citrate 0.6gm
Malachite Green 0.4gm
Magnesium Sulfate 0.24gm

Final pH 7.2 +/- 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 medium to dark green-blue.

Store the prepared culture media at 2-8ºC.

PRECAUTIONS

METHOD OF PREPARATION

1. Suspend 37.2gm of the dehydrated culture media in 600ml of distilled or deionized water containing 12ml of glycerol.

2. Heat to boiling and mix to dissolve completely.

3. Sterilize in the autoclave at 121ºC. for 15 minutes. Cool to 45-60ºC. avoiding air bubbles.

4. Add the sterile base to 1 liter of a uniform suspension of fresh eggs prepared under aseptic conditions. Swirl gently to avoid introducing air into the suspension.

5. Dispense into sterile screw cap tubes or bottles. Arrange in a slanted position.

6. Place in an inspissator, waterbath or autoclave at 85ºC. for 45 minutes to coagulate medium.

PROCEDURE AND INTERPRETATION OF RESULTS

For information on procedures and interpretation of results, consult listed references or refer to the prepared media Instructions for Use (IFU) for Cat. No. C21.

LIMITATIONS

Lowenstein Jensen Media require incubation in a 5-10% CO2 atmosphere in order to recover mycobacteria. Mycobacteria, for unknown reasons, are not recovered well from candle extinction jars.(4)

Protect the media from all sources of light, as malachite green is very photosensitive.

Selective media often inhibit, to some extent, specific strains of organisms for which they are designed to select.

The color of LJ Media may range from a pale-green to a dark blue-green. Do not use media that has turned yellow, as it will interfere with the interpretation of the pigmentation of mycobacteria. Most contaminating bacteria will turn the media blue.

Negative culture results do not rule out an active mycobacterial infection. Some factors responsible for unsuccessful cultures are:

1. The specimen was not representative of the infectious material (For example, saliva instead of sputum).

2. The mycobacteria were destroyed during digestion and decontamination of the specimen.

3. Gross contamination interfered with the growth of mycobacteria.

MATERIALS REQUIRED BUT NOT PROVIDED

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

QUALITY CONTROL

Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Mycobacterium tuberculosis
H3RV
ATCC ® 27294

G 21 days 35°C CO 2 ** Growth; colonies seen in 2 weeks, mature in 3 weeks
Mycobacterium kansasii
Group I
ATCC ® 12478

G 21 days 35°C CO2** Growth; colonies seen in 2 weeks, mature in 3 weeks
Mycobacterium scrofulaceum
Group II
ATCC ® 19981

G 21 days 35°C CO 2 ** Growth; colonies seen in 2 weeks, mature in 3 weeks
Mycobacterium intracellulare
Group III
ATCC ® 13950

G 21 days 35°C CO 2 ** Growth; colonies seen in 2 weeks, mature in 3 weeks
Mycobacterium fortuitum
Group IV
ATCC ® 6841

G 21 days 35°C CO2 ** Growth; colonies visible in 4 days

User Quality Control

PHYSICAL APPEARANCE

CRITERION™ Lowenstein Medium Base powder should appear homogeneous, free-flowing, and in medium to dark green-blue in color. The prepared media should appear opaque, and pale green in color.

REFERENCES

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

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

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

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

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

6. Lowenstein, E. 1933. Ann. Inst. Pasteur.; 50:161.

7. Lowenstein, E. 1931. Zentralbl. Bakteriol. Parasetenkd. Infektionskr.; 120:127.

8. Jensen, F. 1932. Zentralbl. Bakteriol. Parasetenkd. Infektionskr.; Abt. 1 Orig., 125:222.


ATCC is a registered trademark of the American Type Culture Collection.

060616gr