Download Cold Acid Fast Stain Technique

Cold Acid Fast Stain Technique
1. Make smear, air dry, heat fix.
2. Cover the smear with a strip of blotting paper.
3. Saturate the paper with Kinyoun’s carbolfuchsin. To avoid crystallization, be sure to periodically
add back drops of the stain. Let the stain remain for 20 minutes.
4. Remove the blotting paper with forceps. Gently rinse with water and drain.
5. Decolorize with acid-alcohol (not the Gram Stain decolorizer) using 1 “healthy” squirt, followed
by a gentle water rinse.
6. Counterstain with methylene blue for 1 minute.
7. Rinse, blot dry with bibulous paper and observe using low power, followed by high dry and then
the oil immersion lens.
Note: This stain is designed to reveal members of the genus Mycobacterium. They are Acid
Fast positive due to a waxy substance in their cell walls known as mycolic acid which holds on to
the carbolfuchsin. All other bacteria will stain Acid Fast negative.
Acid Fast positive cells are stained the pink/red color of carbolfuchsin.
Acid Fast negative cells are stained the light blue color of methylene blue.