Stain used in preparation of AFB smear:

Ans. (c) Ziehl-neelsen stainRef: Microbiology by Ananthanarayan and Paniker 8th ed. / 352* Z-N stain is used to distinguish acid fast bacteria and non-acid fast bacteria.* Presence of mycolic acid is responsible for the acid-fast nature of bacteria.* For the diagnosis and control of tuberculosis Sputum microscopy is the most reliable single method.* Preparation of smear is from the thick purulent part of the sputum. They are the dried, heated fixed and stained by the Ziehl-Neelsen technique.* The smear is then covered with strong carbol fuchsin and gently heated for 5-7 minutes, without letting the stain boil and become dry.* Wash the slide with water and decolourise it with 20% sulphuric acid, followed by 95% ethanol for two minutes.* Smear is then counterstained with Loeffler s methylene blue, 1% picric acid or 0.2% malachite green for one minute.* Now, under the oil immersion objective, acid fast bacilli are seen as bright red rods while the background is blue, yellow or green depending on the counterstain used.* At least 10,000 acid fast bacilli should be present per ml of sputum for them to be readily demonstrable in direct smears.