Ziehl -- Neelsen stain - wikipedia Ziehl -- Neelsen stain This article needs additional citations for verification. Please help improve this article by adding citations to reliable sources. Unsourced material may be challenged and removed. ( April 2011 ) Mycobacterium tuberculosis visualization using the Ziehl -- Neelsen stain. Ziehl - Neelsen ( Acid - Fast Stain ) diagram of basic steps The Ziehl -- Neelsen stain, also known as the acid - fast stain, was first described by two German doctors : the bacteriologist Franz Ziehl ( 1859 -- 1926 ) and the pathologist Friedrich Neelsen ( 1854 -- 1898 ). It is a special bacteriological stain used to identify acid - fast organisms, mainly Mycobacteria. Mycobacterium tuberculosis is the most important of this group because it is responsible for tuberculosis ( TB ). Other important Mycobacterium species involved in human disease are Mycobacterium leprae, Mycobacterium kansasii, Mycobacterium marinum, Mycobacterium bovis, Mycobacterium africanum and members of the Mycobacterium avium complex. Acid - fast organisms like Mycobacterium contain large amounts of lipid substances within their cell walls called mycolic acids. These acids resist staining by ordinary methods such as a Gram stain. It can also be used to stain a few other bacteria, such as Nocardia. The reagents used are Ziehl -- Neelsen carbol fuchsin, acid alcohol, and methylene blue. Acid - fast bacilli will be bright red after staining. A variation on this staining method is used in mycology to differentially stain acid - fast incrustations in the cuticular hyphae of certain species of fungi in the genus Russula. It is also useful in the identification of some protozoa, namely Cryptosporidium and Isospora. The Ziehl -- Neelsen stain can also hinder diagnosis in the case of paragonimiasis because the eggs in an ovum and parasite sputum sample ( OnP ) can be dissolved by the stain, and is often used in this clinical setting because signs and symptoms of paragonimiasis closely resemble those of TB. Contents 1 Procedure 2 Mechanism explanation 3 Modifications 4 See also 5 References 5.1 Online protocol examples 6 References 7 External links Procedure A typical AFB stain procedure involves dropping the cells in suspension onto a slide, then air drying the liquid and heat fixing the cells. The slide is flooded with Carbol Fuchsin, which is then heated to dry and rinsed off in tap water. The slide is then flooded with a 1 % solution of hydrochloric acid in isopropyl alcohol ( or methanol ) to remove the carbol fuchsin, thus removing the stain from cells that are unprotected by a waxy lipid layer. Thereafter, the cells are stained in methylene blue and viewed on a microscope under oil immersion. Studies have shown that an AFB stain without a culture has a poor negative predictive value. An AFB Culture should be performed along with an AFB stain ; this has a much higher negative predictive value. Mechanism of acid fast staining in acid fast cells and non-acid fast cell. Mechanism explanation Initially, Carbol Fuchsin stains every cell. When they are destained with acid - alcohol, only non-acid - fast bacteria get destained since they do not have a thick, waxy lipid layer like acid - fast bacteria. When counter stain is applied, non-acid - fast bacteria pick it up and become blue when viewed under the microscope. Acid - fast bacteria retain Carbol Fuchsin so they appear red. Modifications 1 % sulfuric acid alcohol for actinomycetes, nocardia. 0.5 -- 1 % sulfuric acid alcohol for oocysts of isospora, cyclospora. 0.25 -- 0.5 % sulfuric acid alcohol for bacterial endospores. Brucella differential stain -- glacial acetic acid used, no heat applied, secondary stain is loeffler 's methylene blue. Kinyoun modification ( or cold Ziehl -- Neelsen technique ) is also available. A protocol in which a detergent is substituted for the highly toxic phenol in the fuchsin staining solution. See also Lowenstein -- Jensen medium References `` Microbiology with Diseases by Body System '', Robert W. Bauman, 2009, Pearson Education, Inc. Morello, Josephine A., Paul A. Granato, Marion E. Wilson, and Verna Morton. Laboratory Manual and Workbook in Microbiology : Applications to Patient Car. 10th ed. Boston : McGraw - Hill Higher Education, 2006. Print. Online protocol examples Ziehl -- Neelsen protocol ( PDF format ). References Jump up ^ Morello, Josephine A., Paul A. Granato, Marion E. Wilson, and Verna Morton. Laboratory Manual and Workbook in Microbiology : Applications to Patient Care. 10th ed. Boston : McGraw - Hill Higher Education, 2006. Print. Jump up ^ Romagnesi, H. ( 1967 ). Les Russules d'Europe et d'Afrique du Nord. Bordas. ISBN 0 - 934454 - 87 - 6. Jump up ^ Largent, D ; D Johnson ; R Watling ( 1977 ). How to identify fungi to genus III : microscopic features. Mad River Press. ISBN 0 - 916422 - 09 - 7. p 25. Jump up ^ `` Online Microbiology Notes ''. Online Microbiology Notes. Retrieved 2017 - 11 - 29. Jump up ^ `` Home - microbeonline ''. microbeonline. Retrieved 2017 - 11 - 29. Jump up ^ Kumar, Surinder ( 2012 ). Textbook of Microbiology. p. 315. Jump up ^ Ellis, RC ; LA Zabrowarny. ( 1993 ). `` Safer staining method for acid fast bacilli '' ( PDF ). Journal of Clinical Pathology. 46 : 559 -- 560. doi : 10.1136 / jcp. 46.6. 559. PMC 501296. PMID 7687254. External links Media related to Ziehl - Neelsen stain at Wikimedia Commons Microbial and histological stains Iron / hemosiderin Prussian blue Lipids Sudan stain Sudan II Sudan III Sudan IV Oil Red O Sudan Black B Carbohydrates Periodic acid -- Schiff stain Amyloid Congo red Bacteria Gram stain Methyl violet / Gentian violet Safranin Ziehl -- Neelsen stain / acid - fast Carbol fuchsin / Fuchsine Methylene blue Auramine - rhodamine stain Auramine O Rhodamine B Connective tissue trichrome stain : Masson 's trichrome stain / Lillie 's trichrome Light Green SF yellowish Biebrich scarlet Phosphomolybdic acid Fast Green FCF Van Gieson 's stain Other H&E stain Haematoxylin Eosin Y Silver stain Grocott 's methenamine silver stain Warthin -- Starry stain Methyl blue Wright 's stain Giemsa stain Gömöri trichrome stain Neutral red Janus Green B Alcian blue stain Movat 's stain Jaswant Singh -- Bhattacharji ( JSB ) stain Tissue stainability Acidophilic Basophilic Chromophobic Retrieved from `` https://en.wikipedia.org/w/index.php?title=Ziehl -- Neelsen_stain&oldid = 842375540 '' Categories : Microscopy Microbiology techniques Laboratory techniques Histopathology Histotechnology Staining dyes Staining Bacteriology Acid fast bacilli Hidden categories : Articles needing additional references from April 2011 All articles needing additional references Talk Contents About Wikipedia Беларуская Български Català Čeština Deutsch Español Euskara Français Galego Italiano עברית Bahasa Melayu Nederlands 日本 語 Polski Português Română Русский Suomi Türkçe Українська 13 more Edit links This page was last edited on 22 May 2018, at 01 : 46. 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Question: what is the secondary stain in the acid fast stain
Answer: loeffler's methylene blue