THE VIRTUAL EDGE: Lab 3 Bacterial Staining Techniques II
Acidfast Stain: Background and Introduction
Mycobacterium and many Nocardia species are called acid-fast because during an acid-fast staining procedure they retain the primary dye carbol fuchsin despite decolorization with the powerful solvent acid-alcohol. Nearly all other genera of bacteria are nonacid-fast. The acid-fast genera have lipoidal mycolic acid in their cell walls. It is assumed that mycolic acid prevents acid-alcohol from decolorizing protoplasm. The acid-fast stain is a differential stain.
Ziehl Neelsen Acid-fast stain
Step 2: Smear Preparation (Review)
Cover the smear with carbolfuchsin dye. Carbolfuchsin a potential carcinogen. Please wear gloves and work with the stain in the hood.
Place a piece of paper towel on top of the dye. Be sure the paper towel is saturated with the dye.
Dry heat for 2 minutes.
Cool and rinse with water.
Decolorize with acid-alcohol for 15-20 seconds.
Wash the top and bottom of slide with water and clean the slide bottom well.
Counterstain with Methylene Blue for 30 seconds to 1 minute.
Wash and blot the slide with bibulous paper.
Focus 10X - then use oil immersion. (Oil immersion review)
Rachel Watson, M.S.
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