kinyoun stain

kinyoun stain


The Kinyoun stain is a fundamental technique in microbiology, used to identify acid-fast bacteria like Mycobacterium tuberculosis and Mycobacterium leprae. This staining method is based on the Ziehl-Neelsen stain recommended by the WHO.

Principles of Kinyoun Stain

 The Kinyoun stain’s principle lies in its use of carbolfuchsin solution, which contains carbolfuchsin and phenol. Acid-fast bacteria possess a unique cell wall with a lipoid capsule, making them challenging to stain. Carbolfuchsin stains this capsule, rendering it resistant to decolorization by acid-alcohol. Consequently, acid-fast bacteria retain their red color, distinguishing them from other microorganisms that absorb the counter-blue stain.

Main Components of the Staining Kit

  1. Carbolfuchsin Solution: This contains carbolfuchsin and phenol, essential for staining acid-fast bacteria.
  2. Acid Alcohol Solution: Comprising ethanol and hydrochloric acid, this solution is crucial for decolorization.
  3. Methylene Blue Solution: With methylene blue as its core component, this solution is used for the final staining step.

Sample Requirement

To prepare a sample for Kinyoun staining, select sputum specimens with cheese-like, pus-like, or suspicious characteristics. Apply approximately 0.05ml of the specimen evenly onto a 10mmx20mm oval sputum on a glass slide. Air dry the specimen, then heat-fix the slide by passing it over an open flame four times, each lasting 5 seconds.

Test Procedure

  1. After fixation, place the smear on a rack, ensuring a 10mm slide spacing.
  2. Apply carbolfuchsin solution at room temperature for 10 minutes or more.
  3. Gently wash one end of the slide to remove excess dye.
  4. Cover the slide with acid alcohol solution for 1-2 minutes for decolorization. Rinse with water if needed.
  5. Rinse the slide with running water for 10-20 seconds.
  6. Apply methylene blue solution for 30-60 seconds.
  7. Rinse the slide gently with running water before microscopic examination.

Result Interpretation

Under a microscope against a light blue background, acid-fast bacteria, like Mycobacterium tuberculosis, appear red, while other bacteria and cells appear blue. Mycobacterium tuberculosis typically exhibits rod-shaped or slightly curved cells with varying dimensions. In well-stained specimens, darker-colored particles may be observed. Fresh specimens may show single bacteria or clusters with a branched arrangement.

Method Limitation

The Kinyoun stain is specifically designed for use with acid-fast bacteria.


  1. Prevent dye from drying on the slide during staining.
  2. Ensure the sputum membrane’s thickness is suitable.
  3. Avoid cedarwood oil, as it may damage the oil lens.
  4. Be cautious of variations in red shades within acid-fast bacteria.
  5. Preserve smears with absorbent paper and gentle wiping.
  6. Adjust staining time in colder temperatures.
  7. Promptly cover reagents after use to prevent volatilization.
  8. Store reagents away from extreme temperatures and direct sunlight.
  9. Use the Kinyoun stain under professional supervision.
  10. Adhere to instructions and practice personal hygiene.
  11. Dispose of waste per guidelines.
  12. Check the packaging for production and expiry dates.
Kinyoun Stain

Kinyoun stain: Crucial in microbiology for detecting acid-fast bacteria, including Mycobacterium tuberculosis and Mycobacterium leprae.

Product Brand: Baso

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