AFB (ZN Stain) 3 consecutive Sample Test Cost & Procedure

Ziehl-Neelsen acid-fast dyeing procedure is utilized to smudge Mycobacterium species. Identification of AFB in dyed and acid-washed smudges investigated microscopically may furnish the initial bacteriologic proof of the existence of mycobacteria in a clinical sample. Stain microscopy is a rapid and comfortable strategy that can be accomplished.

There are two strategies generally manipulated for acid-fast staining:

Carbolfuchsin strategies which comprise the Ziehl-Neelsen and Kinyoun methods

Fluorochrome strategy utilizing auramine-O or auramine-rhodamine stains

The precept of the Ziehl-Neelsen Method of Acid-Fast Staining

Mycobacteria, which do not smudge generously by Gram dye, are smudged with carbol fuchsin integrated with phenol.

• In the ‘hot’ ZN procedure, the phenol-carbol fuchsin pigment is heated to facilitate the pigment to infiltrate the waxy mycobacterial compartment wall.
• In the ‘cold’ procedure known as Kinyoun Method, smudges are not heated but the penetration is accomplished by boosting the amount of essential fuchsin and phenol and incorporating a ‘wetting mechanism’ chemical.

Exhibition & Preparation

Due to the overnight proliferation of discharges, first morning samples are better likely to generate reasonable healing of AFB.

Immediate Smear: Smear set instantly from a patient sample before processing.

Indirect Smear:  Smear set from a processed sample after centrifugation

Reagents needed:

• Carbol fuchsin stain
• Acid alcohol 3% v/v
• Malachite green 5 g/l

Ziehl-Neelsen Staining Procedure

• Formulate the sputum smudge
• Scatter the sputum evenly over the prominent area of the slip using a constant rotational activity. The instructed extent of the smudge is about twenty mm by ten mm.
• Spot glides on the dryer with smeared texture upwards, and air dry for approximately half an hour.
• Warmth bribe parch smear.
• Warmth fascination with untreated samples may NOT kill M. tuberculosis whereas liquor fascination is bactericidal.
• Application of a preliminary pigment
• Coat the smudge with carbol fuchsin pigment
• Warm the smudge until the moisture just starts to grow.
• Do not overheat. Add different stains if crucial. Allow the heated pigment to stay on the slide for five minutes.
• Wash off the pigment with sterile water.
• Application of decolorizer
• Coat the smudge with three % v/v acid alcohol for two-five minutes or until the smudge is adequately decolorized, i.e. pale pink. Inspection to notice that no additional red pigment runs off the surface when the slip is pitched. Adding a bit of additional decolorizer for very dense glides or those that resume to “bleed” red stain.
• Wash nicely with sanitary moisture
• Coat the pigment with a malachite green dye for one-two minute.
• Wash off the pigment with sterile water