Formula

Pancreactic Digest of Casein, 15.0 g

Agar, 15.0 g

Soy Peptone, 5.0 g

Sodium Chloride, 5.0 g

Yeast Extract, 1.0 g

Dextrose, 1.0 g

Sodium Fumarate, 1.0 g

Sodium Lactate, 0.6 g

Sodium Succinate, 0.5 g

Sodium Formate, 0.5 g

Potassium Nitrate, 0.5 g

Sodium Carbonate, 0.4 g

L-Cysteine, 0.4 g

Menadione, 1.0 mg

Hemin (0.1%Soln), 1.0 ml

Diluted Laked Sheep Blood, 30.0 ml

Sheep Serum, 40.0 ml

Distilled Water, 930.0 ml

 

Final pH 7.5 +/- 0.3 at 25 degrees C.

Final weight 20.0 g +/- 2.0 for AS-546.

 

Precautions

For IN VITRO DIAGNOSTIC USE only.  Approved biohazard precautions and aseptic techniques should be observed when using this product.  This product is for use only by properly-trained and qualified personnel.  Sterilize all biohazard waste prior to disposal.

Storage and Shelf Life

Storage:  Upon receipt, store at room (13°C-27°C) temperature in original container until use.  Avoid overheating or freezing.  Do not use medium if there are signs of deterioration (shrinking, cracking or discoloration due to oxidation of media) or contamination.

The expiration date applies to the product in its original packaging and stored as directed.  Do not use product past the expiration date shown on the container.

Shelf Life:  90 days from date of manufacture.

 

Procedure

Specimen Collection:  Specimens for anaerobic culture should be protected from air (oxygen) during collection, transport and processing.  Consult appropriate references for detailed instructions concerning collection and transport of anaerobes.

Methods for Use:  ETSA should be inoculated directly with clinical material or a broth that has been previously inoculated from clinical material.  Inoculated plates should be streaked to obtain isolated colonies, immediately placed in an anaerobic atmosphere and incubated at 35-37^oC for 18 – 48 hours.  Additional periods of incubation may be necessary to recover some anaerobes.  Detailed instructions for processing anaerobic cultures can be found in the appropriate references.

 

Materials Required But Not Provided

Standard microbiological supplies and equipment such as loops, saline blanks, slides, staining supplies, microscope, incinerator or autoclave, incubators, anaerobic chamber or anaerobic jars, disinfectant, other culture media and serological and biochemical reagents.

 

Limitations

ETSA will not provide complete information for identification of bacterial isolates.  Additional test procedures and media are required for complete identification.  Consult reference materials for additional information.

 

Quality Control

This medium should support good growth of many fastidious and non-fastidious anaerobes isolated from clinical specimens.  It should also facilitate pigmentation of the black pigmenting Prevotella and Porphyromonas spp.

The following organisms are routinely used for quality assurance performance testing at Anaerobe Systems. 

Organism Tested	ATCC #	Results	Time (Hours)	Special Reaction
Bacteroides fragilis	25285	Growth	24 h
Prevotella melaninogenica	25845	Growth	24-48 h	Pigment*
Fusobacterium necrophorum	25286	Growth	24-48 h
Fusobacterium nucleatum	25586	Growth	24 h
Clostridium perfringens	13124	Growth	24 h
Peptostreptococcus anaerobius	27337	Growth	24 h
Prevotella intermedia	25611	Growth	24 h	Pigment*
Propionibacterium acnes	6919	Growth	24-48 h
Eubacterium lentum	43055	Growth	24 h
Actinomyces viscosus	43146	Growth	24-48 h
Porphyromonas gingivalis	33277	Growth	24 h	Pigment*

                                 * Pigment production may require more than 48 hours of incubation.

User Quality Control: The final determination to the extent and quantity of user laboratory quality control must be determined by the end user.

If sterility testing is to be performed on this product, the appropriate percentage of the original shipment amount should be incubated anaerobically and aerobically for 48 – 96 hours.

If the nutritive capacity of this medium is to be tested for performance, it is recommended that the following ATCC organisms be evaluated for growth.

 

Organism	Expected Growth	Special Reaction
B. fragilis	24 hrs
P. melaninogenica	24-48 hrs	pigment
F. nucleatum	24 hrs
C. perfringens	24 hrs
P. anaerobius	24 hrs
P. intermedia	24-48 hrs	pigment
P. gingivalis	24-48 hrs	pigment

 

Physical Appearance:  ETSA should appear opaque to translucent and amber in color in a 15mm x 100mm over-fill sized plate (AS-546).

 

References

1.        Dowell, V. R., Jr. and T. M. Hawkins.  1974.  Laboratory Methods in Anaerobic Bacteriology.  CDC Laboratory Manual.  USDHEW C. D. C. Atlanta, GA 30333.

 

2.        Holdeman, L. V., F. P. Cato, and W. E. C. Moore.  1977.  Anaerobe Laboratory Manual.  Virginia Polytechnic Institute and State University. Blacksburg, VA 24061.

 

3.        Engelkirk, P. G., Duben-Engelkirk, J. and Dowell, V. R.  1992.  Principles and Practices of Clinical Anaerobic Bacteriology.  Star Publishing Co., Belmont, CA 94002.

 

4.        Jousimies-Somer, H. R., Summanen, P., Citron D. M., Baron, E. J., Wexler, H. M. and S. M. Finegold.  2002.  Wadsworth – KTL Anaerobic Bacteriology Manual.  Star Publishing Co., Belmont, CA 94002.

 

5.        NCCLS.  Quality Control for Commercially Prepared Microbiological Culture Media; Approved Standard- Third Edition.  (2004).  NCCLS document M22-A3.  NCCLS, 940 West Valley Road, Suite 1400, Wayne, PA 19087-1898.