3.10: Acid-Fast Stain

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Page ID: 90987

Rosanna Hartline
Lemoore College

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Preparation

Have test tubes with sterile TSB on hand.
12-18 hours before the laboratory, inoculate test tubes of sterile TSB with a mixture of Staphylococcus epidermidis and Mycobacterium chelonae and incubate at 37 °C .

Materials

12-18 hour cultures (enough for groups to share)
plain microscope slides (at least 1 per group)
Bunsen burners (1 per group)
inoculating loops (1 per group)
(if using bacteria on solid medium rather than TSB) droppers with saline, DI water, or distilled water
slide warmer
test tube racks (1 per group plus one or more racks to hold cultures)
400-500 mL beakers with ~200 mL of water each (1 per group)
hot plate (1 per group)
slide rack hat fits over beaker (1 per group)
paper towels
carbol fuchsin in dropper bottles (1 per group)
acid-alcohol in dropper bottles (not ethanol from the Gram stain kit) (1 per group)
methylene blue in dropper bottles (1 per group)
wood test tube clamps/holders (1 per group)
small chemical waste containers (1 per group)
distilled or DI water in squirt bottles (1 per group)
bibulous paper (enough for each groups to blot their slides)
light microscopes (1 per group)
colored pencils
(optional, but recommended) Wax pencils (1 per group)
(if using oil immersion) immersion oil dropper bottles
(if using oil immersion) lens paper

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