The Significance of Acid-Fast Stain


Different types of bacterial species have certain properties that make them unique from one another. Scientists use a special technique called staining to help them distinguish one bacterial species from another when observed under light microscopy. One staining technique often used to identify pathogenic (disease-causing) bacteria is acid-fast staining.


  • German bacteriologist Franz Ziehl discovered the affinity of the bacterial genus Mycobacterium for the red dye carbolfuchsin in 1882. He found when these particular bacteria were flooded with the dye, nothing would decolorize them, not even acids. This is where scientists get the term acid-fast.


  • All members of the bacterial genus Mycobacterium are acid-fast. This is of significant clinical importance because of two specific species in this genus: Mycobacterium tuberculosis and Mycobacterium leprae, the causative agents for tuberculosis and leprosy, respectively.

Carbolfuchsin Affinity

  • Mycobacterium and Norcardia both contain a special waxy material called mycolic acid in their cell walls. When carbolfuchsin dye is poured over these cells, it becomes so deeply embedded inside them it cannot be removed. This is because carbolfuchsin is more soluble in the lipids of the cell wall than in the acid-alcohol.

Staining Procedure

  • The acid-fast staining procedure is simple. First, a small drop of water is placed on a microscope slide, and a bacterial sample is smeared into the water drop. The water air-dries, and carbolfuchsin dye is poured over the entire slide. Once this is done, the slide is gently heated for several minutes. Next, the slide is allowed to cool and then rinsed heavily with water. This removes any excess dye. Next, acid-alcohol is poured over the slide. This removes any carbolfuchsin dye from any non-acid fast bacteria. Finally, methylene blue stain is poured over the slide to stain any remaining non-acid-fast bacteria.


  • The ability to identify the causative agents of tuberculosis and leprosy using acid-fast staining techniques cannot be underestimated. Even though the discovery of this staining technique dates back to 1882, it remains an important part of helping to maintain the public health.


  • Microbiology: An Introduction; Gerard Tortora, Berdell Funke, Christine Case; 2010
  • Acid Fast Stain
  • Photo Credit Microscope image by e-pyton from
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