Acid fast staining

Acid fast staining (AFS) is a laboratory tests that is found by scientists Franz Ziehl and Friedrich Neelsen and also called Ziehl-Neelsen staining. This laboratory test is commonly used to identify mycobacteria and some kinds of other microorganisms. These microorganisms have a property of acid resistance because of the lipid in their cell walls and called acid fast bacteria (AFB). Mycobacterium Tuberculosis is also a acid-fast bacterium and can be identified with acid-fast staining method.

Purpose of Acid-Fast Staining

Quick diagnosis; acid fast staining method provides quick results and it is a cheap and easy method so may be preferred to diagnose tuberculosis.

Confirming the bacteria’s acid fast property that are obtained from culture test.

Monitoring the treatment is going well or not.

Acid Fast Staining Samples

Any kind of clinical material and the microorgansims that is provided from mycobacterial culture can be used for acid fast staining For exp.

acid fast staining

  • Blood
  • Urine
  • Stool
  • Bone marrow
  • Tissue
  • Sputum
  • Body fluids
  • Fluids from joints
  • Cerebrospinal fluid (CSF)


ImportantTopics About Acid Fast Staining

All sample preparation process must be done in the biosafety cabinet. Biosafety cabinet protects the performers health and prevents contamination to environment. Results must be controlled with negative and positive control smears.

How the Acid Fast Staining Test is Performed

First decontamination process is done to the sample to eliminate the bacteria other than the acid-fast bacteria (AFB).

After decontamination process a little amount of sample is taken from the sediment to make a smear on a slide. Smear may be about 1,5×1,5 cm big same as a coin.

Smear may be done without gaining a sediment from decontamination but this will reduce acid fast bacteria determination chance.

After smear preparation slides are waited for air drying, then slides will be ready for acid fast staining.

Carbolfuchsin, acid alcohol and methylene blue are used for acid fast staining in an order.

1- Carbolfuchsin

AFB and the other bacteria will be red-purplish

2- Acid-alcohol

AFB remain red but the others release the dye, this is decolorization

3- Methylene blue

AFB remain red again but others take the dye inside and become blue.

Evaluation of Acid Fast Staining Samples

Before resulting the smaple negative (clear) at least 300 fields must be analysed under microscope. If there are no bacteria in these 300 fields result will be negative but before writing the test report the other slides must be analysed too.

  • If only 1-2 bacteria in 300 fields test must be repeated
  • 1-9 bacteria in 100 fields 1+
  • 1-9 bacteria in 10 fields 2+
  • 1-9 bacteria in 1 field 3+
  • More than 10 bacteria in every fields 4+

Acid fast staining result must be reported in 24 hours.


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