ACID FAST STAINING
Kinyoun staining (Cold Method)
1. What is Acid fast staining?
Property of the cell wall of certain bacteria, their structures, protozoal forms by
vertue of which they resist decolorisation by mineral acids following staining with
strong acids like carbol fuchsin.
2. Name a few acid fast organisms?
Mycobacterium tuberculosis
Mycobacterium leprae
Nocardia
Oocyst of Cryptosporidium, Cyclospora, Isospora
Record the observations of acid fast organisms
M. leprae
M.tuberculosis
Nocardia Coccidian parasites
, 3. What is the principle of Acid fast staining ?
Since the waxy mycoloic acid containing cell wall of mycobacteria is
relatively impermiable to ordinary staining techniques. Staining by aniline
dye is possible either by application of heat that softenes the wax in the cell
wall or by using solvents such as phenol
The basic fuchsin dye is soluble in phenol. The dye phenol mixture enter
the cells.
Once stained it resist decolorisation by mineral acids such as 25% H 2So4.
While the mycobacteria retains the primary dye, the background material
gets decolorised and takes up the counter stain.
4. What are the components of Acid fast staining?
Primary stain : Concentrated carbol fushion with phenol
Decolourization agent : 25% sulphuric acid
Counter stain : Methylene blue .
5. What are the different methods of Acid fast staining ?
1. Heat method or Zeihl Neelsen method 2. Cold method or Kinyon’s method
6. What is the procedure for kinyon’s method?
1. Identify the correct side of the slide to be stained
2. Flood the slide with concentrated carbol fuchsin (with phenol) and leave it for 7 mins
3. Wash it with water and add Gabbett’s methylene blue
(which contains loeffler’s methylene blue and 25 % of H2So4)and leave it for
3mins
4. Wash the slide, air dry it and observe under oil immersion (100X)
Kinyoun staining (Cold Method)
1. What is Acid fast staining?
Property of the cell wall of certain bacteria, their structures, protozoal forms by
vertue of which they resist decolorisation by mineral acids following staining with
strong acids like carbol fuchsin.
2. Name a few acid fast organisms?
Mycobacterium tuberculosis
Mycobacterium leprae
Nocardia
Oocyst of Cryptosporidium, Cyclospora, Isospora
Record the observations of acid fast organisms
M. leprae
M.tuberculosis
Nocardia Coccidian parasites
, 3. What is the principle of Acid fast staining ?
Since the waxy mycoloic acid containing cell wall of mycobacteria is
relatively impermiable to ordinary staining techniques. Staining by aniline
dye is possible either by application of heat that softenes the wax in the cell
wall or by using solvents such as phenol
The basic fuchsin dye is soluble in phenol. The dye phenol mixture enter
the cells.
Once stained it resist decolorisation by mineral acids such as 25% H 2So4.
While the mycobacteria retains the primary dye, the background material
gets decolorised and takes up the counter stain.
4. What are the components of Acid fast staining?
Primary stain : Concentrated carbol fushion with phenol
Decolourization agent : 25% sulphuric acid
Counter stain : Methylene blue .
5. What are the different methods of Acid fast staining ?
1. Heat method or Zeihl Neelsen method 2. Cold method or Kinyon’s method
6. What is the procedure for kinyon’s method?
1. Identify the correct side of the slide to be stained
2. Flood the slide with concentrated carbol fuchsin (with phenol) and leave it for 7 mins
3. Wash it with water and add Gabbett’s methylene blue
(which contains loeffler’s methylene blue and 25 % of H2So4)and leave it for
3mins
4. Wash the slide, air dry it and observe under oil immersion (100X)
