Commons is a freely licensed media file repository. The main aim of this staining is to differentiate bacteria into acid fast group and non acid fast groups. The current classification of staining procedures for tb microscopy was made based on the. Acidfast stain mycobacterium and many nocardia species are called acidfast because during an acidfast staining procedure they retain the primary dye carbol fuchsin despite decolorization with the powerful solvent acidalcohol 95% ethanol with 3% hcl. The evaluator must understand appropriate methods and ensure they are being followed. Acid fast staining differentiates acid fast bacteria from non acid fast bacteria based on the mycolic acid content in the cell wall. The acid fast stain robert koch was the first person to isolate and identify mycobacterium tuberculosis from a patient with tuberculosis. Acid fast staining in formalinfixed tissue specimen of. Make a thin film of the material on a clean glass slide, using a sterile loop or swab for viscous specimens. The acidfast genera have lipoidal mycolic acid in their cell walls. The oil hastens acid fast staining without the use of heat.
Non acid fast bacteria quickly lose the colour of the primary stain due to the absence of mycolic acid and appears blue. Nowadays, ziehlneelsen method is believed as most important differential staining procedure used for the identification of acid fast species of mycobacterium. This staining technique is used to demonstrate the presence of acid fast bacilli. Acid fast bacteria include the mycobacteria and some of the nocardia. Analysis of specimens for the detection of acid fast bacilli by microscopy. Acid fast bacilli red leprae bacilli are short rods nocardia red nocardia organisms are long, thin, and filamentous erythrocytes pale pink. Online microbiology notes welcome to, constantly growing and evolving collection of microbiology notes and information. If you continue browsing the site, you agree to the use of cookies on this website. They can be stained by aniline dyes using drastic measures such as application of heat and. There are certain bacteria which cannot be stained by grams method.
Its application aim at read more albert staining principle, reagents, procedure, results, interpretation. Capsule is synthesized in the cytoplasm and secreted to the outside of the cell where it surrounds the bacterium. The evaluator needs to have a good understanding of acidfast bacilli afb smear microscopy and good laboratory practice glp. Paul ehrlich is the first to describe the acidfast properties of the bacterium. Acidfast stain principle, procedure, interpretation and. Methods assessed include specimen collection, smear preparation, acid fast stain procedure, reading, and reporting. It is named for two german doctors who modified the stain. Jul 28, 2017 acid fast staining differentiates acid fast bacteria from non acid fast bacteria based on the mycolic acid content in the cell wall. The purpose of afb microscopy is to detect acidfast bacilli afb by microscopic. The acid fast bacterial cell envelope is a specialized derivation of the grampositive cell envelope that has an extremely high lipid content. Most common of the acid fast staining techniques is the ziehlneelsen technique.
Difference between gram stain and acid fast compare the. Acid fast stain is for the demonstration of ber cu lo sismycobacterium tu in tissue sections and smears. Ziehlneelsen n stain acid fast stain linkedin slideshare. Acid fast mycobacteria contain mycolic acid in their outer membrane, making the cells waxy and resistant to staining with aqueous based stains such as the gram stain.
This method was named, and still known as acidfast staining and the bacteria were named as acidfast bacteria. Standard operating procedure sop for hematoxylin and eosin. Acid fast is a stain used for bacteria that have a particularly waxy cell wall. Feb 28, 2018 z n stain is a modification of ehrlichs original method for differential staining of acid fast bacilli by use of aniline gentian violet followed by strong nitric acid.
Nearly all other genera of bacteria are nonacid fast. Gram positive microorganisms that are resistant to gram stain, acid fast allows us to see this bacteria mycolic acid a waxy substance in cell walls of acid fast bacteria. Examine under high dry magnification and verified under oil immersion. Paul ehrlich is the first to describe the acid fast properties of the bacterium. Such cells are thus termed acidfast in that the cell will hold the stain fast in the presence of the acidic decolorizing agent. This procedure uses pararosaniline dye in a phenol solution. Detection of acid fast bacilli afb in stained and acid washed smears examined microscopically may provide the initial bacteriologic evidence of the. It can also be used to stain a few other bacteria, such as nocardia. Steamers are very hot, and acid alcohol is highly flammable. The primary stain, carbolfuchsin is applied to the cells, and heat and phenol are used to allow the stain to penetrate into the waxy surface of acid fast microorganisms. View the article pdf and any associated supplements and figures for a period of 48 hours. Acid fast organisms like mycobacterium contain large amounts of lipid substances within their cell walls called mycolic acids. This method is used for those microorganisms which are not staining by simple or gram staining method, particularly the member of genus mycobacterium, are resistant and can only be visualized by acidfast staining. In the acid fast staining procedure, mycobacteria are first stained with the red dye basic fuchsin and the cells are then decolorized and restained with methylene blue.
Partially acid fast organisms show reddish to purple filaments as opposed to non acid fast organisms, which stain only blue. Sep 11, 2009 acid fast stainingsummary microbiology lab 11th septravi a patel slideshare uses cookies to improve functionality and performance, and to provide you with relevant advertising. Acid fast staining is another commonly used, differential staining technique that can be an important diagnostic tool. A cold staining method for acidfast bacilli article pdf available in bulletin of the world health organisation 645. The primary stain, carbolfuchsin, is applied to the cells and phenol is used to allow the stain to penetrate into the waxy surface of acidfast microorganisms. Acid fast stainingsummary microbiology lab 11th septravi a patel slideshare uses cookies to improve functionality and performance, and to provide you with relevant advertising. The second is the lightemitting diode led that has a life span of more than 50,000 hours, is less expensive, and has a performance similar to the mvpbased microscopy. This property is possessed by few bacteria other than mycobacterium. Nonacidfast bacteria are counterstained with methylene blue. The evaluator needs to have a good understanding of acid fast bacilli afb smear microscopy and good laboratory practice glp.
This method is used for those microorganisms which are not staining by simple or gram staining method, particularly the member of genus mycobacterium, are resistant and can only be visualized by acid fast staining. Acidfast staining and microscopic examination are the easiest, quickest, and least expensive diagnostic procedures. Acidfast bacteria include the mycobacteria and some of the nocardia. Most of the capsulated bacteria have a capsule made. Acidfast mycobacteria contain mycolic acid in their outer membrane, making the cells waxy and resistant to staining with aqueous based stains such as the gram stain. If washing with distilled water is not done adequately, crystallization of the stain may appear on the slide. Pdf on oct 30, 2018, sandeep sharma and others published acid fast bacteria find, read and cite all. Each of these stains aims at identifying and characterizing bacteria based on their morphologies. In the method described here longer exposure favours the uniformpenetration ofthe dyethrough thecellwall. The most common stain being gram staining, acidfast staining, endospore staining. He developed a stain for the bacterium, although it was not very effective for visualizing this slender bacillus. Stain with carbol fuchsin for 20 minutes at room temperature.
This document describes the standard operating procedure for microscopic analysis for. There should be a thin film of oil on the slide at all times during the procedure. So by use of powerful staining solution that contain phenol and application of heat, the dye. Manual of histopathological and histochemical procedures, 16th ed. Detection of acidfast bacilli afb in stained and acidwashed smears examined microscopically may provide the initial bacteriologic evidence of the presence of mycobacteria. The main aim of this staining is to differentiate bacteria into acid fast group and nonacid fast groups. It is the differential staining techniques which was first developed by ziehl. This is the difference between acid fast and gram stain. Moreover, this hypothesis is indirectly supported by reports that the fite method, which is an alternative acidfast staining technique that uses less xylene in the staining and mounting process, has a higher sensitivity than ziehlneelsen staining 7, 8. Because of this feature, this stain is extremely helpful in identification in diseases caused by acid fast bacteria, particularly tuberculosis and leprosy.
This file is licensed under the creative commons attributionshare alike 4. In the acidfast staining procedure, mycobacteria are first stained with the red dye basic fuchsin and the cells are then decolorized and restained with methylene blue. These acids resist staining by ordinary methods such as a gram stain. Carbolfuchsin acid fast stain is used to demonstrate mycobacterium tu ber cu lo sis in tissue sections and on smears. The stained smear is the easiest and quickest procedure that can be performed for diagnosis of tb and is economical especially in developing countries. The waxy mycolic acid containing cell wall of mycobacteia are relatively impermeable to ordinary staining techniques. Mycolic acid is a waxy substance that does not allow the decolourizer to enter into the cell wall due to its waxy nature. Acid fast stain is for the demonstration of mycobacterium tu ber cu lo sis in tissue sections. Fluorochrome staining an overview sciencedirect topics. Sensitivity of acidfast staining for mycobacterium. Standard operating procedure sop for hematoxylin and. Principle, procedure, reporting and modifications by editorial team on january 11, 2020 in bacteriology, microbiology the ziehlneelsen stain zn stain, also called the hot method of afb staining, is a type of differential bacteriological stain used to identify acidfast organisms, mainly mycobacteria.
Information from its description page there is shown below. An acid fast stain is able to differentiate two types of grampositive cells. Acid fast stain kinyouncold method principle, procedure. Ziehlneelsen zn method of acid fast staining technique is used to stain mycobacterium species including m.
Acknowledgments uk standards for microbiology investigations uk smis are developed under the auspices of public health england phe working in partnership with the national. Coa is not included with the antibiotic shipment, it can be downloaded by lot number from. Acid fast stain principle, reagents, procedure and result. Jun 10, 2018 this method was named, and still known as acid fast staining and the bacteria were named as acid fast bacteria. The zn stain confirms the acidfast property of mycobacteria. Endospore stain procedure moeller and schaefferfulton methods duration. You might see these terms used interchangeably or see staining reagents labelled with zn rather than af. Basic fuchsin binds to lipids in the cell wall and turns mycobacteria red while other bacteria stain blue. Mycobacterium and many nocardia species are called acidfast because during an acidfast staining procedure they retain the primary dye carbol fuchsin despite decolorization with the powerful solvent acidalcohol. Whether youre a student, professor, working in the medical field or just curious about microbiology, im sure youll find our articles interesting and informative. The primary stain, carbolfuchsin, is applied to the cells and phenol is used to allow the stain to penetrate into the waxy surface of acid fast microorganisms.
Ziehlneelsen staining is a bacteriological stain used to identify acidfast organisms, mainly mycobacteria. The acidfast stain robert koch was the first person to isolate and identify mycobacterium tuberculosis from a patient with tuberculosis. Choose from 500 different sets of acid fast staining flashcards on quizlet. Therefore, acidfast staining is used to differentiate the mycobacterium species from the other groups of bacteria. In addition, the stain is used to determine the presence of af bacteria from.
Use millipore filtered water in the waterbath and staining procedure. Read download file report abuse differential staining of bacteria. Acidfast stain principle, procedure, interpretation and examples. Jun 12, 2018 the main aim of this staining is to differentiate bacteria into acid fast group and non acid fast groups. Acidfast stain definition of acidfast stain by medical. Acid fast stain mycobacterium and many nocardia species are called acid fast because during an acid fast staining procedure they retain the primary dye carbol fuchsin despite decolorization with the powerful solvent acid alcohol 95% ethanol with 3% hcl. The acid fast staining property results from the presence of membrane glycolipids and very long chain 2alkyl3hydroxy fatty acids. The ziehlneelsen method for staining acidfast bacteria. Avoid overstaining as some weak staining bacterial staining can be replaced by methylene blue.
Studying on the granules of acidfast bacilli observed by using the various staining procedures, the author found the following results. Acid fast stain reagents are for in vitro diagnostic use. Steamers are very hot, and acidalcohol is highly flammable. In mycobacteria, up to 60% of the dry weight of the cell envelope consists of lipid.
After staining, it is essential that the back surface of the slide is wiped clean. In the same year, ehrlichs method was improved by zehil and neelsen. Nearly all other genera of bacteria are nonacidfast. This document details the lab protocol for nhanes 20032004 data. This procedure is performed by trained histology laboratory personnel. Carbolfuchsin solution, microwave 80 power, 45 seconds, allow slides to stand in hot solution. Microscopic examination of tissues or of sputum stained by the acidfast staining procedure is an aid in the diagnosis of tuberculosis.
The oil hastens acidfast staining without the use of heat. Mycobacteriology laboratory manual world health organization. Decolorize in 1% acid alcohol until the slide is a faint pink. The acidfast staining property results from the presence of membrane glycolipids and very long chain 2alkyl3hydroxy fatty acids mycolic acids bound to the peptidoglycan.
Sep 14, 2016 the ziehlneelsen method for staining acid fast bacteria. Learn acid fast staining with free interactive flashcards. In 1882, paul ehrlich developed a method of staining such type of bacteria. Acid fastness is an uncommon characteristic shared by the genera mycobacterium and nocardia weakly acid fast. Z n stain is a modification of ehrlichs original method for differential staining of acid fast bacilli by use of aniline gentian violet followed by strong nitric acid.
The ordinary aniline dye solution do not readily penetrate the acid fast bacilli. Endospore stain procedure moeller and schaefferfulton methods. After staining with basic fuchsin, acidfast bacteria resist decolonization by acidalcohol. Jan 11, 2020 acid fast mycobacteria contain mycolic acid in their outer membrane, making the cells waxy and resistant to staining with aqueous based stains such as the gram stain. Ziehlneelsen staining is a type of acidfast stain, first introduced by paul ehrlich. Methods assessed include specimen collection, smear preparation, acidfast stain procedure, reading, and reporting. Ziehlneelsen acidfast staining procedure medical laboratories.
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