An acidfast stain is a laboratory test performed on a sample of. Its main application is in the staining and demonstration of acid fast bacilli in sputum smears. Microorganisms that are not acidfast nonacid fast will appear blue due to the counterstaining with methylene blue after they have been decolorized by the acid alchol. Nearly all other genera of bacteria are nonacidfast. Your doctor can order this test to find out if you have tuberculosis tb or another type of bacterial. Past history of pulmonary tuberculosis was positive in 46% of patients. Once stained as part of a sample, these organisms can resist the acid andor ethanolbased decolorization procedures common in many staining protocols, hence the name acidfast. The waxy mycolic acid containing cell wall of mycobacteia are relatively impermeable to ordinary staining techniques. They can be stained by aniline dyes using drastic measures such as application of heat and. Microscopic examination of tissues or of sputum stained by the acid fast staining procedure is an aid in the diagnosis of tuberculosis. Most bacterial species are either gram positive or gram negative, however some organisms have different cell wall properties. It is also grampositive, but the stain is weak so it is much better to use acid fast staining. The most commonly known acid fast staining technique is the ziehlneelsen technique. Acid fast bacterium an overview sciencedirect topics.
There are 2 different methods of acidfast stainingboth involve techniques that make the cell wall more permeable to the primary stain. By colouring otherwise transparent tissue sections, these stains allow highly trained pathologists and researchers to view, under a microscope, tissue morphology structure or to look for the presence or prevalence of particular cell types, structures or even microorganisms such as bacteria. Mycobacterium species resist the effect of the acid. Structure and composition of the acid fast cell wall.
Fulltext staining techniques for detection of acid fast bacilli. Acid fast bacteria include the mycobacteria and some of the nocardia. The acid fast stain is a differential stain used to identify acid fast organisms such as members of the genus mycobacterium acid fast organisms are characterized by waxlike, nearly impermeable cell walls. Then the cells are washed with a dilute acid alcohol solution. This is adifferential stain used to identify acid fast bacterial organisms, such as members of the genus mycobacterium and nocardia. Comparison of the value of two different sputum staining for. It prefers colder temperatures, and thus sticks to infecting the extremities of a person and the. Dec, 2017 is video me hamne afb stain ke baare me bataya hai. Acid fast stain lab report essays and research papers.
The ordinary aniline dye solution do not readily penetrate the acid fast bacilli. 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 acid alcohol. It is assumed that mycolic acid prevents acidalcohol from decolorizing protoplasm. Ziehlneelsen staining is a type of acidfast stain, first introduced by paul ehrlich. Koch used the method to observe the tubercle bacilluswhat we now call mycobacterium tuberculosis, in sputum samples. Acid fast bacteria are grampositive, but in addition to peptidoglycan, the outer membrane or envelope of the acid fast cell wall of contains large amounts of glycolipids, especially mycolic acids that in the genus mycobacterium, make up approximately 60% of the acid fast cell wall figure \\pageindex2\. While acid fast and gram staining are both differential stains, the acid fast stain is much more specific. Two different methods for acid fast staining are the ziehlneelsen technique and the kinyoun technique. This file is licensed under the creative commons attributionshare alike 4.
Record susceptible, resistant, or test failed on the internal lab worksheetbook and. Acid fastness in the property of certain bacteria mycobacteria, their structures spores or protozoal forms oocyts. In the zn acidfast stain is a fairly vigorous staining procedure using carbolfuchsin karbalfyukshun as the main stain and methylene blue as a counter stain other counter stains may be used but we will ignore them for simplicity. A rapid method for distinction between gramnegative and grampositive bacteria by means of a 3% solution of potassium hydroxide is tested on 71 grampositive and 55 gramnegative bacterial strains. Ziehlneelsen staining was performed for acid fast bacilli on paraffin embedded sections of tissue blocks. Thus, various staining methods are developed to facilitate bacterial detection. Media kit reprints and eprints sponsored supplements branded books. Paul ehrlich is the first to describe the acid fast properties of the bacterium. The ziehlneelsen stain, also known as the acid fast stain, was first described by two german doctors. Acidfast stain principle, procedure, interpretation and. Acid fast stain is a differential stain used to identify acid fast organisms such as members of the genus mycobacterium.
Partially acid fast organisms show reddish to purple filaments as opposed to non acid fast organisms, which stain only blue. An acid fast stain is a laboratory test performed on a sample of. The primary for acid fast is carbolfuchsin dye and the counterstain is methylene blue. Introduction to acid fast and endospore staining most bacterial species are either gram positive or gram negative, however some organisms have different cell wall properties that make them difficult to stain with this method. He developed a stain for the bacterium, although it was not very effective for visualizing this slender bacillus. Neelsen in 1883 used ziehls carbolfuchsin and heat then decolorized with an acid. Jun 12, 2018 it is the differential staining techniques which was first developed by ziehl and later on modified by neelsen. Acidfast stain principle, procedure, interpretation and examples. Acid fast staining was developed in ehrlich in 1882 and was later successively modified by ziehl and neelsen. Compare and contrast the gram stain and the acidfast stain. Acid fast and endospore staining biology libretexts. The microscope is a very important tool in microbiology, but there are limitations when it comes to using one to observe cells in general and bacterial cells in particular.
Nearly all other genera of bacteria are nonacid fast. It is also important to differentiate tuberculous from nontuberculous mycobacterial lymphadenitis because their treatment protocols vary. Lab 5 acid fast stain questionsanswers flashcards quizlet. The acid fast stain robert koch was the first person to isolate and identify mycobacterium tuberculosis from a patient with tuberculosis. Many acid fast bacteria are not stained with the common stain like carbol fuchsin because they are prepared in aqueous solution. The nocardia are aerobic, grampositive rods and stain partially acid fast i. An intro to routine and special staining in histopathology. Acidfast organisms are characterized by waxlike, nearly impermeable cell walls. Your health care provider will collect a sample of urine, stool, sputum, bone marrow, or tissue, depending on the location of the suspected infection. Get a printable copy pdf file of the complete article 809k, or click on a page image below to browse page by page. It was devised by a danish physician, hans christian gram, in 1884. In the ziehlneelsen stain, a fixed smear covered with carbolfuchsin is heated, rinsed, decolorized with acid alcohol, and counterstained with methylene blue.
The technique is also referred to as acid fast staining. Learn quiz microbiology lab acid fast staining with free interactive flashcards. The detection limit of an acidfast stain has been estimated to be 500010 000 bacilliml of sputum. Two acidfast genera that are important as human pathogens are mycobacterium and nocardia. Start studying lab 5 acid fast stain questionsanswers. Key difference gram stain vs acid fast bacteria are very small microorganisms. This staining technique was discovered by scientist paul ehrlich in 1883. The mechanisms of acidfastness vary by species, although the most wellknown example is in the genus myc. So by use of powerful staining solution that contain phenol and application of heat, the dye.
Functions of reagents used in acid fast staining technique. Differential staining techniques viewing bacterial cells. Ziehlneelsen staining is a bacteriological stain used to identify acidfast organisms, mainly mycobacteria. Acidfastness is a physical property of certain bacterial and eukaryotic cells, as well as some subcellular structures, specifically their resistance to decolorization by acids during laboratory staining procedures. Acid fuchsin is a magenta red acid dye that is largely used for plasma staining whereas basic fuchsin is a magenta basic dye largely used to stain the nucleus. The acid fast bacterial cell envelope is a specialized derivation of the grampositive cell envelope that has an extremely high lipid content. Cell wall treated to allow penetration of the dye either by heat ziehlneelsen method or by a detergent kinyoun method. Mycobacteriology laboratory manual world health organization. Another differential stain technique is the acid fast technique. The manual covers lab safety, how to use a microscope, aseptic transfer of bacteria, simple, gram, capsule, acid fast, spore, and negative staining, environmental requirements of bacteria, antibiotic susceptibility testing, bacterial control methods, membrane filtration, standard plate count, plaque assay, biochemical tests for the. Technical variations in both the preparation of clinical material and subsequent staining mean that smear sensitivity relative. The most common site of involvement was pleura followed by vertebral and lymph nodes. Most common of the acid fast staining techniques is the ziehlneelsen technique.
The purpose of afb microscopy is to detect acidfast bacilli afb by microscopic. The acid fast staining property results from the presence of membrane glycolipids and very long chain 2alkyl3hydroxy fatty acids. It is a special bacteriological stain used to identify acid fast organisms, mainly mycobacteria. Soon after kochs discovery, paul ehrlich developed a stain for. Heat or a lipid solvent is used to carry the first stain, carbolfuchsin, into the cells. Full text full text is available as a scanned copy of the original print version. Ziehlneelsen n stain acid fast stain linkedin slideshare. The nature of the acidfast stain pubmed central pmc. Comparison of conventional ziehlneelsen method of acid. Links to pubmed are also available for selected references. Acid fastness is a physical property of certain bacterial and eukaryotic cells, as well as some subcellular structures, specifically their resistance to decolorization by acids during laboratory staining procedures. One acidfast staining method for mycobacterium tuberculosis is the ziehlneelsen stain. The cell walls of acid fast organisms contain a wax like lipid called mycolic acid. Such cells are thus termed acid fast in that the cell will hold the stain fast in the presence of the acidic decolorizing agent.
Kappa statistics revealed good agreement between results of both pap staining and ar staining method and culture k 0. 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. Detailed instruction on staining by ziehl neelsen method. There are no serological tests, and the drug of choice is. Todars online textbook of bacteriology chapter on tuberculosis tb, the most deadly bacterial disease in history. Acid fast smear evaluation and reportinga report no. This book is distributed under the terms of the creative commons. Gram stain, a widely used microbiological staining technique that greatly aids in the identification and characterization of bacteria. If you want to know if something is an acid, you can test it by using litmus paper. This property is possessed by few bacteria other than mycobacterium. Acidified alcohol is used as a decolorizer, and methlyene blue is the counterstain. Grampositive acid fast nocardia brasiliensis bacteria using a modified fitefaraco stain. So this method is also called ziehlneelsen staining techniques.
The acid fast stain is a laboratory test that determines if a sample of tissue, blood, or other body substance is infected with the bacteria that causes tuberculosis tb and other illnesses. Two of the most important concerns are resolution and contrast. Acidfast staining technique helps us to differentiate the organism as acidfast and nonacid fast organisms. Chemistry of neutralization and salt formation introduction. As a result the cell wall is relatively impermeable to ordinary staining techniques. Staining techniques book summaries, test preparation. The term acid fast bacilli is practically synonymous with mycobacteria, although nocardia and some other organisms are variably acid fast. Choose from 500 different sets of quiz microbiology lab acid fast staining flashcards on quizlet. Staining solutions for acid fast staining sigmaaldrich. The acid fast stain is a first step in identification of these types of bacteria. Acid fast staining was developed by robert koch in 1882 and later modified by other scientists. Ziehlneelsen staining is a type of acid fast stain, first introduced by paul ehrlich. Acid fast staining in tuberculosis linkedin slideshare.
They are transparent, and their detection is difficult under living and unstained conditions. A variety of staining techniques can be used with light microscopy, including gram staining, acid fast staining, capsule staining, endospore staining, and flagella staining. Is video me hamne afb stain ke baare me bataya hai. Detection of acid fast bacilli in saliva using papanicolaou. For staining such organism ziehlneelsen staining method is used. Apr 17, 2017 history of acid fast staining in 1882 robert koch reported the discovery of the tubercle bacillus and described the appearance of the bacilli resulting from a complex staining procedure. Theoretically, there are 2 staining methods of tb microscopy that were. Acid fast staining in formalinfixed tissue specimen of.
This may involve staining, fixation, andor cutting thin sections. The acidfast stain is a differential stain used to identify acidfast organisms such as members of the genus mycobacterium acidfast organisms are characterized by waxlike, nearly impermeable cell walls. Introduction to acid fast and endospore staining biology. 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. Once stained as part of a sample, these organisms can resist the acid andor ethanolbased decolorization procedures common in many staining protocols, hence the name acid fast.
Acidfast stain definition of acidfast stain by medical. The zn stain confirms the acidfast property of mycobacteria. Alternative staining techniques kinyoun or acid fast stain are therefore used that take advantage of the resistance to. Microbiology acid fast staining technique and its details. Cell staining in microscopy types, techniques, preparations. Neelsen in 1883 used ziehls carbolfuchsin and heat then decolorized with an acid alcohol, and counter stained with methylene blue. Acidfast stain is a differential stain used to identify acidfast organisms such as members of the genus mycobacterium. Learn vocabulary, terms, and more with flashcards, games, and other study tools.
Sputum for acid fast bacilli afb is seldom looked for in the etiological diagnosis of tuberculous pleural effusion usually due to the absence of any parenchymal lesion radiologically, but presence of tubercle bacilli in sputum may have important epidemiological and therapeutic implication. The material sent to the lab, depending on the presentation of the disease, is sputum, pus, or biopsy material. Acid fast organisms are characterized by waxlike, nearly impermeable cell walls. Role of sputum examination for acid fast bacilli in. An acid fast stain is able to differentiate two types of grampositive cells. Once stained as part of a sample, these organisms can resist the acid andor. A photomicrograph of mycobacterium smegmatis pink and micrococcus luteus blue at x magnification. The acidfast stain robert koch was the first person to isolate and identify mycobacterium tuberculosis from a patient with tuberculosis. Terms in this set 10 what is the primary and counter stain for acid fast stain. Ziehlneelsen staining is a bacteriological stain used to identify acid fast organisms, mainly mycobacteria. Acids will turn litmus paper red, whilst alkalis will turn it blue. It is the differential staining techniques which was first developed by ziehl and later on modified by neelsen. Furthermore, an acidfast stain can quickly assess patient infectiousness, and is also used to confirm that a culture is acid fast.
Franz ziehl 1859 to 1926, a bacteriologist and friedrich neelsen 1854 to 1894, a pathologist. Acidfast bacteria also known as acidfast bacilli or afb are microorganisms resistant. Mar 05, 20 acid fast af is an important special staining technique used in the histology lab. Despite its long history, the acid fast smear remains unstandardised. A thorough history and physical examination, staining for acid fast bacilli afb, and polymerase chain reaction pcr are helpful in obtaining an early diagnosis. Acidfast staining questions and study guide quizlet. Acid fast microorganisms will retain the dye of carbolfuchsin and appear red. During the same time period several other researchers ehrlich, ziehl, rindfleisch, and neelsen, intending to improve on kochs method, introduced. Certain bacteria and actinomycetes have many components in the cell wall and their cell wall has little permeability. 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. Paul ehrlich was one of those rare individuals in the history of medicine who made. The acidfast genera have lipoidal mycolic acid in their cell walls.
The gram reaction reflects fundamental differences in the biochemical and structural properties of bacteria. It is named for two german doctors who modified the stain. In the united states an estimated 500 1,000 new cases of nocardiosis infection occur annually. In the ziehlneelsen stain, a fixed smear covered with carbolfuchsin is heated, rinsed, decolorized with acid. Acid fast retain primary stain red, others dont blue. For example, some species of bacteria have a waxy lipid mycolic acid in their cell walls. Cvi crystal violet iodine complex complex is formed. Many bacteria are either gram positive or gram negative, but very few are acidfast. Acids are positively charged ions, they are liquid and are solutions of pure compounds in water. Examine under high dry magnification and verified under oil immersion. The protozoan parasite cryptosporidium can also be stained using this procedure. Acidfast staining technique is a differential staining technique in bacteriology. Aug 20, 2011 acidfast stain how it works acidfast cells contain a large amount of lipids and waxes in their cell walls primarily mycolic acid acid fast bacteria are usually members of the genus mycobacterium or nocardia therefore, this stain is important to identify mycobacterium or nocardiadr.