Ex. 5: Acid-Fast and Endospore Stains

Objectives:Explain the rationale and procedure for the acid-fast and endospore stainsPerform and interpret the acid fast and endospore stainsReview the structure and function of endospores

For Acid Fast Bacteria

Use only

Carbol fuchsin for AFB!!

Acid Alcohol

MethyleneBlue

Modified Kinyoun’s Acid-fast Stain – Cold Method

• Heat fix smear• Primary stain: Carbol fuchsin for AFB, (5 min)• Rinse with water• Decolorize briefly (~10 drops) with acid alcohol

(~ 30 sec)• Immediately rinse with water• Counterstain: Methylene blue (1 min)• Rinse with water

Endospore Stain

• Heat fix smear• Primary stain: Malachite green, steam (5

min)• Rinse thoroughly with water (~ 30 sec)• Counterstain: Safranin (1 min)