Acid-Fast Staining

Acid-fast staining, a differential staining procedure, is also known as Ziehl-Neelsen method as it was first developed by Franz Ziehl in 1882 and later modified by Friedrich Neelsen in 1883. Some bacteria are surrounded by a covering of fatty and waxy substances and are difficult to stain. But once they are stained, they can’t be easily decolorized even after the treatment with alcohol containing acid. Such organisms, because of their resistance to decolorization with acid alcohol are known as acid-fast. A few species particularly those in the genus Mycobacterium are acid-fast like Mycobacterium tuberculosis (causing tuberculosis) and Mycobacterium leprae (causative agent of leprosy).

In this staining procedure, cells are first heated with a primary stain i.e. Carbol fuchsin. This will stain every cell as red. Then they are destained with an acid-alcohol. Acid-fast cells have high lipid content in particular mycolic acid-a group of branched chain hydroxyl lipids so they can’t be easily decolorized even by treatment with alcohol and hence stains red. Non-acid fast bacteria lack the thick, waxy lipid layer like that of acid-fast bacteria and hence they lose the primary stain and get destained by acid wash. Finally, counter stain i.e. methylene blue is applied. Therefore acid-fast bacteria appears red in color and non acid-fast bacteria appears blue in color when viewed under the microscope.