The Kinyoun method or Kinyoun stain (cold method), developed by

Joseph J. Kinyoun Joseph James Kinyoun (November 25, 1860 – February 14, 1919) was an American physician and the founder of the United States' Hygienic Laboratory, the predecessor of the National Institutes of Health. Biography Early life Joseph James "Jo ...

, is a procedure used to

stain A stain is a discoloration that can be clearly distinguished from the surface, material, or medium it is found upon. They are caused by the chemical or physical interaction of two dissimilar materials. Accidental staining may make materials app ...

acid-fast Acid-fastness is a physical property of certain bacterial and eukaryotic cells, as well as some sub-cellular structures, specifically their resistance to decolorization by acids during laboratory staining procedures. Once stained as part of a sam ...

species of the bacterial genus ''

Mycobacterium ''Mycobacterium'' is a genus of over 190 species in the phylum Actinomycetota, assigned its own family, Mycobacteriaceae. This genus includes pathogens known to cause serious diseases in mammals, including tuberculosis ('' M. tuberculosis'') and ...

''. It is a variation of a method developed by

Robert Koch Heinrich Hermann Robert Koch ( , ; 11 December 1843 – 27 May 1910) was a German physician and microbiologist. As the discoverer of the specific causative agents of deadly infectious diseases including tuberculosis, cholera (though the Vibrio ...

in 1882. Certain species of bacteria have a waxy lipid called mycolic acid, in their cell walls which allow them to be stained with Acid-Fast better than a Gram-Stain. The unique ability of mycobacteria to resist decolorization by acid-alcohol is why they are termed acid-fast. It involves the application of a primary stain (

basic fuchsin Fuchsine (sometimes spelled fuchsin) or rosaniline hydrochloride is a magenta dye with chemical formula C20H19N3·HCl.

), a decolorizer (acid-alcohol), and a counterstain (

methylene blue Methylthioninium chloride, commonly called methylene blue, is a salt used as a dye and as a medication. Methylene blue is a thiazine dye. As a medication, it is mainly used to treat methemoglobinemia by converting the ferric iron in hemoglobin ...

). Unlike the

Ziehl–Neelsen stain Ziehl–Neelsen staining is a type of acid-fast stain, first introduced by Paul Ehrlich. Ziehl–Neelsen staining is a bacteriological stain used to identify acid-fast organisms, mainly Mycobacteria. It is named for two German doctors who modif ...

(Z-N stain), the Kinyoun method of staining does not require heating. In the Ziehl–Neelsen stain, heat acts as a physical mordant while phenol (carbol of carbol fuschin) acts as the chemical mordant. Since the Kinyoun stain is a cold method (no heat applied), the concentration of carbol fuschin used is increased.

Staining procedure

Kinyoun Stain, organism Mycobacterium smegmatis

Make an acid-fast stain https://faculty.mtsac.edu/cbriggs/Micr-22NewLabmanual2Aug2018.pdf

Materials

* Slide with organism smear * Carbol Fuchsin * Acid-Alcohol * Methylene blue * bibulous paper * Microscope

Instructions

# Make smear on a slide with organisms #* Clean slide, wax label slide, spread organism, air dry for 10 minutes, heat fix # Dip slide into Carbol Fuchsin for 20 minutes. # Rinse slide # Dip slide into Acid-alcohol for 3–5 seconds. # Rinse Slide # Dip slide into Methylene blue for 30 seconds. # Rinse Slide # Blot slide dry with bibulous paper. # Observe under Microscope.

Modification

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''.

References