Download Laboratory 6- Acid Fast Stain

BIOL 2120
HCC-Stafford Campus
J.L. Marshall, Ph.D.
Laboratory 6- Acid Fast Stain*
*Laboratory notes are to be used as a study guide only and do not represent the comprehensive information you will need to know
for the exams.
The acid fast stain is a differential stain that tests for the presence of mycolic acids, a thick waxy layer,
associated with the cell wall of acid fast bacteria.
Medically important Acid fast bacteria –
1. Mycobacterium species – i. M. tuberculosis (tuberculosis) and, ii. M. leprae (leprosy, or Hansen’s
disease) are two of the more common pathogenic species of this genus.
2. Nocardia species – N. asteroids, an opportunistic pathogen.
Cell structure:
Acid fast
Gram positive
mycolic acids in cell wall
cell wall
cell membrane
cell membrane
mycolic acids: a waxy layer on the outer surface of the acid fast bacterium, used to protect them against
hostile environments. Affects staining, a barrier to most dyes. Since they are difficult to destain, even with
acid, they are called acid fast.
The Acid fast stain procedure:
~ prepare an air dried, heat fixed smears of the bacterial culture
Step 1: primary stain, carbolfuchsin
-
all cells stain red
Step 3: decolorize with acid-alcohol
- non acid fast = clear
- acid fast = red
Step 2: modant, place slide over boiling water
-
all cells will be red
Step 4: counter stain with methylene blue
- non acid fast = blue
- acid fast = red
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BIOL 2120
HCC-Stafford Campus
J.L. Marshall, Ph.D.
The acid fast stain is a differential stain. In the Gram stain acid fast bacteria stain as Gram positive, so you have
to perform an acid fast stain to differentiate between the acid fast bacteria and the Gram positive bacteria.
Gram stain
Gram negative
Gram positive (true?)
Acid fast stain
Non-acid fast (blue)
Acid fast (red)
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