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Ziehl–Neelsen stain (classic and modified bleach types) [5]; Kinyoun stain; For color blind people (or in backgrounds where detecting red bacteria is difficult), Victoria blue can be substituted for carbol fuchsin and picric acid can be used as the counter stain instead of methylene blue, and the rest of the Kinyoun technique can be used.
Mechanism of acid-fast staining in acid-fast cells and non-acid-fast cell [24] [25] [26] The mechanism of action of the Ziehl-Neelsen stain is not completely understood, but it is thought to involve a chemical reaction between the acidic dyes and the cell walls of the bacteria.
The Kinyoun method can be modified as a weak acid fast stain, which uses 0.5–1.0% sulfuric acid instead of hydrochloric acid.The weak acid fast stain, in addition to staining Mycobacteria, will also stain organisms that are not able to maintain the carbol fuchsin after decolorizing with HCl, such as Nocardia species and Cryptosporidium.
A Ziehl–Neelsen stain is an acid-fast stain used to stain species of Mycobacterium tuberculosis that do not stain with the standard laboratory staining procedures such as Gram staining. This stain is performed through the use of both red coloured carbol fuchsin that stains the bacteria and a counter stain such as methylene blue .
Some bacteria have cell walls which are particularly adept at retaining stains. These will appear positive by Gram stain even though they are not closely related to other gram-positive bacteria. These are called acid-fast bacteria, and can only be differentiated from other gram-positive bacteria by special staining procedures. [25]
[2] [3] Carbol fuchsin is used as the primary stain dye to detect acid-fast bacteria because it is more soluble in the cells' wall lipids than in the acid alcohol. If the bacteria is acid-fast the bacteria will retain the initial red color of the dye because they are able to resist the destaining by acid alcohol (0.4–1% HCl in 70% EtOH). [4 ...
Mycobacterium smegmatis is an acid-fast bacterial species in the phylum Actinomycetota and the genus Mycobacterium. It is 3.0 to 5.0 μm long with a bacillus shape and can be stained by Ziehl–Neelsen method and the auramine-rhodamine fluorescent method.
Gram staining uses two dyes: Crystal violet and Fuchsin or Safranin (the counterstain) to differentiate between Gram-positive bacteria (large Peptidoglycan layer on outer surface of cell) and Gram-negative bacteria. Acid-fast stains are also differential stains.