What is the procedure of ZN staining?
Ziehl Neelsen Acid-fast stain
- Step 2: Smear Preparation (Review)
- Cover the smear with carbolfuchsin dye.
- Dry heat for 2 minutes.
- Cool and rinse with water.
- Wash the top and bottom of slide with water and clean the slide bottom well.
- Counterstain with Methylene Blue for 30 seconds to 1 minute.
What is ZN stain test?
Conventional smear microscopy with the Ziehl-Neelsen (ZN) stain is a rapid and practical method for detecting acid-fast bacilli (AFB), especially in low-income countries, due to its rapidity, low cost, and high positive predictive value for tuberculosis (14).
What is the critical step in ZN staining?
The critical step of this staining process is the decolorizing step, which can be influenced by the thickness of the specimen smear. In addition, a fresh solution of hydrochloric acid in ethanol is highly reac- tive, meaning that the result should be evaluated with caution.
What are the 3 main steps of an acid-fast stain?
Acid-Fast Staining Instructions
- Air dry and heat fix a thin film of microorganisms.
- Flood the slide with Carbolfuchsin.
- Flood slide with Acid Alcohol for 30 seconds.
- Counterstain by flooding the slide with Methylene Blue for 30 seconds.
- Dry the slide by putting it between the pages of a book of Bibulous paper.
What is ZN stain in microbiology?
Ziehl–Neelsen staining is a bacteriological stain used to identify acid-fast organisms, mainly Mycobacteria. It is named for two German doctors who modified the stain: the bacteriologist Franz Ziehl (1859–1926) and the pathologist Friedrich Neelsen (1854–1898).
What is the principle of AFB?
The mycobacterial cell wall contains mycolic acids, which are fatty acids that contribute to the characteristic of “acid-fastness.” The principle of the AFB smear is based on the fact that mycolic acid in the cell wall of AFB render them resistant to decolorization with acid alcohol.
What is Zn technique?
Ziehl-Neelsen (ZN) method of acid-fast staining technique is used to stain Mycobacterium species including M. tuberculosis, M. ulcerans, and M. leprae and nontuberculous mycobacteria (NTM).
What are the procedural steps of Ziehl-neelsen?
Ziehl-Neelsen Staining Procedure
- Prepare the sputum smear.
- Place slides on the dryer with smeared surface upwards, and air dry for about 30 minutes.
- Heat fix dried smear.
- Application of a primary stain.
- Heat the smear until vapor just begins to rise (i.e. about 60°C).
- Wash off the stain with clean water.
Why 20% H2SO4 is used in ZN staining?
The waxy material is hydrophobic to aqueous solution but not to phenolic solution of basic fuchsin. Hence strong carbol fuchsin is able to stain the cell. Upon staining, they tend to resist decolourization by 20% H2SO4 (sulphuric acid).
Why Sulphuric acid is used in ZN staining?
Which bacteria are acid-fast?
Acid-fast bacteria, also known as acid-fast bacilli or simply AFB, are a group of bacteria sharing the characteristic of acid fastness….These include:
- Bacterial endospores.
- Head of sperm.
- Cryptosporidium parvum.
- Isospora belli.
- Cyclospora cayetanensis.
- Taenia saginata eggs.
- Hydatid cysts.
- Sarcocystis.
Why do we use Zn test for some bacteria?
Microscopic examination of clinical samples for acid-fast bacilli using the Ziehl-Neelsen (ZN) stain has been a standard diagnostic tool and is used globally for rapid TB diagnosis. Microscopy can detect 60% to 70% of culture-positive samples with a lower limit of detection of 5 × 103 organisms/mL.
Why is acid alcohol used in acid-fast staining?
Acid alcohol has the ability to completely decolorize all non-acid-fast organisms, thus only leaving behind red-colored acid-fast organisms, like M. tuberculosis. The slides are then stained a second time with methylene blue that serves as a counterstain.
Is TB gram-positive or negative?
tuberculosis belongs to the high G+C Gram-positive bacteria that form a monophyletic group with the low G+C Gram-positive bacteria such as Bacillus subtilis. Some analyses indicate no particular relationship between these two groups.
Why is methylene blue used in acid-fast staining?
After the auramine dye has fully stained the smear, a drop of acid alcohol is applied for one to two minutes to decolorize the smear. Methylene blue or potassium permanganate is used as a counterstain to provide background color.
What is AFB test?
Brief Description. Acid- Fast Bacilli (AFB) smear and culture are two separate tests always performed together at the MSPHL, Tuberculosis (TB) Unit. AFB smear refers to the microscopic examination of a fluorochrome stain of a clinical specimen.
Which acid is used in ZN staining?
sulphuric acid in water for Ziehl-Neelsen staining of acid-fast bacilli.