|Mycobacterium avium, Ziehl-Neelsen staining
Acid fast organisms have a lipoid capsule that can resist decolorization with a dilute acid rinse (previously stained with Ziehl-Neelsen
fuchsin solution). Method is used for genus Mycobacterium.
Ziehl-Neelsen fuchsin solution: basic fuchsin 2.5 g, distilled water 250 ml, 100%
alcohol 25 ml, phenol crystals melted 12.5 ml.
Acid solution: H2SO4 in H2O 1:4.
Loffler methylene blue solution: methylene blue crystals 1 g, phenol crystals melted,
96% alcohol 10 ml, distilled water 100 ml.
Dry and heat-fix the slide then cover it with Ziehl-Neelsen fuchsin solution. Heat the
slide for 10 minutes; avoid boiling. Wash in running tap water.
Add H2SO4 solution and wait few seconds. Remove acid solution and wash in running
Cover the slide with Loffler methylene blue solution and wait 1 minute. Remove
solution, wash and dry the slide.
Acid-fast bacteria are red colored, others are blue. After discoloring only acid fast cells remain red.
Solutions are flammable and corrosive.
(c) Costin Stoica