ACID FAST STAIN: AURAMINE RHODAMINE

Technical Data #AURAMINEa / 2013.07.21

The Auramine-Rhodamine is used to stain acid-fast organisms as well as Mycobacterium sp. in specimens and in culture. This fluorescent method, actually the best procedure, stains selectively mycobacteria by binding dye to the mycolic acid of the cell wall.

 


 

FORMULA
in grams per liter

Auramine Rhodamine, Stain

 

Auramine O

11,1

Rhodamine B

5,6

Phenol

74,0

Glycerol

556,0 ml

Water, deionized

370,0 ml

Acid-Alcohol, Decolorizer

 

Ethanol, denatured

700,0 ml

Hydrochloric Acid, concentrated

5,0 ml

Water, deionized

300,0 ml

Potassium Permanganate, Counterstain

 

Potassium Permanganate

5,0

Water, deionized

1000,0 ml


PRECAUTIONS
This medium is for IN VITRO diagnostic use only.


STORAGE
Store in tightly-sealed containers at 15-30ºC in the dark.


SIGN OF DETERIORATION
Stains should not be used if they show signs of deterioration or if the expiration date has passed.


PROCEDURE

  1. Prepare the smears as described in previous standardized methods directly from specimens or from culture.
  2. Fix smears by passing through a Bunsen burner for 10 seconds.
  3. Flood slides with Auramine-Rhodamine and stain for 15 minutes.
  4. Rinse thouroughly with tap water and decolorize with Acid-Alcohol for 2 minutes.
  5. Rinse with tap water and counterstain with 0,5% Potassium permanganate for 3 minutes.
  6. Rinse with tap water. Blot dry and examine microscopically under a fluorescent microscope at low and high magnification.


QUALITY CONTROL
 

Organisms

ATCC

Color

Morphology

Escherichia coli

25922

green

bacilli

Mycobacterium tuberculosis

25177

fluorescence

bacilli


LIMITATIONS OF METHOD
Slides of acid-fast should be stained individually to prevent acid fast bacilli carry-over from one slide to another. This is a part of identification. Other methods may be required.

 

REFERENCES

  1. Finegold, S.M., E.J. Baron 1986 Bailey and Scott's Diagnostic Microbiology 7th ed. C.V. Mosby St. Louis
  2. Lennette, E.H., A. Balows, W.J. Hausler Jr., H.J. Shadomy 1985 Manual of Clinical Microbiology 4th ed. American Society for Microbiology, Washington, D.C.
  3. Vestal, A.L., 1969 Procedures for the isolation and identification of mycobacteria. USDHEW, Public Health Service No. 1995, U.S. Government Printing. Washington, D.C.


CATALOGUE NUMBERS

Solutions    
Auramine Rhodamine, Stain 500 ml 8537
Acid-Alcohol, Decolorizer 500 ml 8541
Potassium Permanganate, Counterstain 500 ml 8538
Complete Set    
Auramine-Rhodamine 3x250 ml 8540
Auramine 3x250 ml 8543

 


 

 

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