Kinyoun stain
Encyclopedia
The Kinyoun stain is a method of staining acid-fast
Acid-fast
Acid-fastness is a physical property of certain bacteria, specifically their resistance to decolorization by acids during staining procedures.Acid-fast organisms are difficult to characterize using standard microbiological techniques Acid-fastness is a physical property of certain bacteria,...

 microorganisms, specifically mycobacterium
Mycobacterium
Mycobacterium is a genus of Actinobacteria, given its own family, the Mycobacteriaceae. The genus includes pathogens known to cause serious diseases in mammals, including tuberculosis and leprosy...

. The procedure for Kinyoun staining is similar to the Ziehl-Neelsen stain
Ziehl-Neelsen stain
The Ziehl–Neelsen stain, also known as the acid-fast stain, was first described by two German doctors; Franz Ziehl , a bacteriologist and Friedrich Neelsen , a pathologist. It is a special bacteriological stain used to identify acid-fast organisms, mainly Mycobacteria...

, but does not involve heating the slides being stained.1 The Kinyoun staining method uses carbolfuchsin as a primary stain, followed by decolorization with an acid-alcohol solution and methylene blue
Methylene blue
Methylene blue is a heterocyclic aromatic chemical compound with the molecular formula C16H18N3SCl. It has many uses in a range of different fields, such as biology and chemistry. At room temperature it appears as a solid, odorless, dark green powder, that yields a blue solution when dissolved in...

 as a counterstain. Kinyoun carbolfuschsin has a greater concentration of phenol
Phenol
Phenol, also known as carbolic acid, phenic acid, is an organic compound with the chemical formula C6H5OH. It is a white crystalline solid. The molecule consists of a phenyl , bonded to a hydroxyl group. It is produced on a large scale as a precursor to many materials and useful compounds...

 and basic fuchsin and does not require heating in order to stain properly.1 When viewed under a microscope, a Kinyoun stained slide will show acid-fast organisms as red and nonacid-fast organisms as blue.

Procedure

  1. Flood slides with Kinyoun carbolfuchsin for 5 minutes.
  2. Rinse gently with water until the water flows off clear.
  3. Flood slides with acid-alcohol (3% HCl in ethanol) for 3 minutes.
  4. Rinse gently with water until the water flows off clear.
  5. Flood slides with methylene blue for 3 minutes.
  6. Rinse gently with water until the water flows off clear.
  7. Allow slides to air dry before viewing.

Modifications

A solution of 1% sulfuric acid can be substituted in place of 3% HCl solution. The sulfuric acid solution doesn't decolorize as strongly as the hydrochloric acid. This makes it useful for staining organisms that are weakly acid fast, such as Nocardia
Nocardia
Nocardia is a genus of weakly staining Gram-positive, catalase-positive, rod-shaped bacteria. It forms partially acid-fast beaded branching filaments . It has a total of 85 species. Some species are non-pathogenic while others are responsible for nocardiosis. Nocardia are found worldwide in soil...

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