When stained with a primary stain and fixed by a mordant, some bacteria are able to retain the primary stain by resisting declorization while other get decolorized by decolorizer. Examine the stained smear with the microscope, under oil immersion. Find powerpoint presentations and slides using the power of, find free presentations research about gram staining ppt. An historical discussion of the gram method for staining bacteria. Gram staining procedure animation microbiology principle. Gram stain principle, reagents, procedure, steps, results. The genera actinomyces, arthobacter, corynebacterium, mycobacterium, and propionibacterium have cell. Gram stain protocols american society for microbiology. Gram stain workshop for the laboratory generalist aphl. Explain the principle of the gram stain procedure, including. Gram staining and the steps needed to complete the procedure. Staining is a technique where specimens are stained with specific dyes to create contrast between the specimen and its background.
Textbook of diagnostic microbiology 6th edition pdf free download is an asset to any student. It was discovered by danish scientist and physician hans christain joachin gram in 1884. Jan 27, 2017 principle of grams staining the structure of the organism s cell wall determines whether the organism is gram positive or negative. A free powerpoint ppt presentation displayed as a flash slide show on id. The staining procedure differentiates organisms of the domain bacteria according to cell wall structure. Because the bacteria are colored, not only is their gram stain group identified, but their shape, size, and clumping pattern may be observed. The gram stain procedure distinguishes between gram positive and gram negative groups by coloring these cells red or violet. If you have ever taking a course or you are new the subject, then i just got you a free ticket to download this wonderful book together with other books for free. Difference between gram positive and gram negative cell wall is thicker in a gram positive bacteria. Basic microbiology lab reporting gram staining free download as powerpoint presentation. The gram staining technique differentiates the mixed culture cells into two terms as gram positive bacteria and gram. Principle, procedure, interpretation, examples and.
Gram stain hans christian gram history and principles of gram. General principles for the formation and proliferation of. Gardnella has an unusual gram positive cell wall structure that causes bacteria of this genus to stain gram negative or gram variable sadhu et al 1989. Gram staining technique is the widely used differential staining technique in bacteriology. Of all the different classification systems, the gram stain has withstood the test of time.
What is gram stain test staining procedure, principle and. Flood the smear with grams iodine and allow the iodine to react for about 2 minutes. Grams iodine in gram staining and phenol in ziehl neelsons staining. Describe common modifications to the gram stain procedure. Decolorization step rinse with ethanol until color stops following then rinse with water. Rinse the slide with tap water, tilting the slide slightly to rinse all the stain from the slide. Our first effort uses ebook readers, which have several ease of reading features already built in. The procedure of gram staining requires four basic steps that include first, applying a primary stain of crystal violet to a heat. Gram in 1884 it remains an important and useful technique to this day. Gram positive bacteria stain blue while gram negative stain red.
Gram stain protocol gram staining is a common technique. A primary stain originally anilinegentian violet was used, but this has been replaced by the more stable crystal violet. Access free multiple choice questions on this topic. View and download powerpoint presentations on gram staining ppt. The gram stain procedure enables bacteria to retain color of the stains, based on the differences in the chemical and physical properties of the cell wall. Certain bacteria are stained with aniline dyes, like gentian violet. Explain the principle behind the gram stain procedure. Reagents of gram stain because the gram stain reaction is more widely used and studied than any other stain, it is also the most open to variation in protocol. The gram staining procedure is one of the most basic and important tests performed in the clinical. Pdf mlt 415 lab report gram stain techniques muhamad. The gram staining technique differentiates the mixed culture cells into two terms as gram positive bacteria and gram negative bacteria. Gram staining is an empirical method of differentiating bacterial species into two large groups gram positive and gram negative based on the chemical and physical properties of their cell walls. Several different types of staining procedure are known today which are used to study morphological features of microorganism and their parts table 1.
To a gram positive cell the counter stain stains the cell, but the cells are already so heavily stained by crystal violet that the addition of pink does not change the dark purple color of the cells. The lipoid capsule of the mycobacteria is of such high molecular weight that it is waxy at room temperature and successful penetration by the aqueousbased staining solutions such as gram s is prevented. Gram staining showing gram positive cocci in clusters, and culture of fluid from a vesicle or pustule or from beneath the lifted edges of a crusted plaque of impetigo, are usually sufficient to establish a diagnosis. Principle the gram stain is used to classify bacteria on the basis of their forms, sizes, cellular morphologies, and gram reactions. The cell walls of gram positive bacteria have a thick layer of proteinsugar complexes called peptidoglycan and lipid content is low. Pdf gram staining procedure find, read and cite all the research you need on researchgate. The basic principle of gram staining involves the ability of the bacterial. The gram stain was first used in 1884 by hans christian gram, he was searching for a method that would allow visualization of cocci in tissue sections of lungs of people who were diagnosed and died of pneumonia. This makes the gram stain a valuable diagnostic tool for a medical clinic or lab. What is gram stain test staining procedure, principle. Flood slide with crystal or gentian violet 60 seconds. Microscopic morphology and staining of individual bacteria. Gram staining is initially established by the physician hans christian gram, which was from denmark. When the diagnosis is in question, and gram stain and culture are negative, a skin.
This is applied and stains the gram negatives pink. Gram negative cells have a thin peptidoglycan layer and stain red to pink. Oct 15, 2005 named after hans christian gram who developed the method in 1884, the gram stain allows one to distinguish between gram. Gram staining was firstly introduced by cristian gram in 1883 which used to. A staining method that uses only a single dye that which does not.
If the control slides do not prove satisfactory, the staining procedure is not accepted. Gram positive cell wall contains a thick layer of peptidoglycan with numerous teichoic acid crosslinking which resists the. Gram stain gram stain pure strain cultures to determine whether gram negative or gram positive. Cell wall of gram positive and gram negative bacteria. It allows a large proportion of clinically important bacteria to be classified as either gram positive or negative based on their. Positive and negative control slides should be used every time the staining procedure is performed except for gram staining where positive controls may be enough unless a new batch of stain is made. Principle, procedure, interpretation, examples and animation.
Developed by danish bacteriologist christian gram in 1884, the gram stain is the most fundamental test used to differentiate bacteria. Three introductory chapters cover gramstain procedures, quality control, evaluation, and more. Gram positive bacteria has only the inner plasma membrane and no outer membrane. Ppt gram stain procedure powerpoint presentation free. Named after hans christian gram who developed the method in 1884, the gram stain allows one to distinguish between gram. Grampositive bacteria stain violet due to the presence of a thick layer of peptidoglycan in their cell. Gather all equipment and supplies gram stain microscope slides heat source sterile swab culture or specimen microscope gram stain procedure 2. Aug 12, 2019 the gram stain is fundamental to the phenotypic characterization of bacteria. Doc principles of staining mohammad ehtisham academia. The bulletins published by the station will be sent free to any farmer amjlviltllll for them. Basic microbiology lab reporting gram staining staining. The results of the gram stain are viewed using light microscopy.
Ppt gramstaining download powerpoint presentations. Gram variable staining due to cell wall changes with loss of viability. Pdf on jan 1, 2018, upasana bhumbla published grams staining find, read and cite all the research. Gram positive cells have a thick peptidoglycan layer and stain blue to purple. Staining methods simple staining, negative staining, grams. Nov 03, 20 it includes introduction, history, principle, method of gram staining procedure slideshare uses cookies to improve functionality and performance, and to provide you with relevant advertising. The gram stain procedure distinguishes between gram positive and gram negative groups by. The lipoid capsule of the acidfast organism takes up carbolfuchsin and resists decolorization with a dilute acid rinse. Nov 25, 2019 difference between gram positive and gram negative cell wall is thicker in a gram positive bacteria. This technique differentiates bacteria in 2 groups i.
The use of pure cultures has made possible the identification of the bacterial etiology of many infectious diseases. A primary stain originally anilinegentian violet was used, but. The differences in cell wall composition of gram positive and gram negative bacteria account for the gram staining differences. Textbook of diagnostic microbiology 6th edition pdf free download. Textbook of diagnostic microbiology 6th edition pdf free.
Gram stains from blood cultures and direct specimen sterile body. Gram staining is performed invitro and is used to identify a bacterial organism based on the amount of. He help to distinguish klebsiella pneumonia to pneumococci. Through a series of staining and decolorization steps, organisms in the domain bacteria are differentiated according to cell wall composition.
Although simple stains are useful, they do not reveal details about the bacteria other than morphology and arrangement. Some bacteria, after staining with the gram stain, yield a gram variable pattern. Stain dark purple due to retaining the primary dye called crystal violet in the cell wall. Gram staining is the common, important, and most used differential. Discuss circumstances which may cause unexpected or erroneous gram stain results.
Differentiation in regressive staining differentiation is the removal of washing out of the excess stain until the colour is retained only in the tissue components to be studies. Simple biochemical characteristics such as the ability to ferment a given carbohydrate. Oct 05, 2020 gram staining is a differential staining technique that is used for microscopic examination of bacteria. Ppt gram stain procedure powerpoint presentation free to. Oct 24, 2019 purpose of the gram staining technique. Gram staining detects a fundamental difference in the cell wall composition of bacteria. Gram stain the previous lab introduced simple staining techniques that enable microbiologists to observe the morphological characteristics of bacteria.
The counter stain used in the gram stain is safranin pink. The composition of the cellular wall of gram positive and gram negative bacteria tells the gram staining differences. The principle of gram s stain has remained unchanged but the technique has been modified in keeping with improvements in the qualities of dyes. In short, the process of gram staining comprises the use of a solution of gram iodine or potassium iodide to the cells which are use to stain before with crystal violet or gentian violet. Remove the crystal violet by rinsing gently with iodine. This was developed in 1884 by a danish bacteriologist, hans christian gram. Pace, in neonatal dermatology second edition, 2008 diagnosis. Gram staining was firstly introduced by cristian gram in 1883 which used to differentiate the gram positive microorganisms and. Staining is an auxiliary technique used in microscopic techniques used to enhance the clarity of the microscopic image. When the bacteria is stained with primary stain crystal violet and fixed by the mordant, some of the bacteria are able to retain the primary stain and some are decolorized by alcohol. Briefly centrifuge all small tubes and vials containing the stainsreagents before opening to prevent waste of reagents.
Gram staining is most widely staining technique used in mo examination. Place the slide on the staining rack and flood the slide with stain for 1minute. If you continue browsing the site, you agree to the use of cookies on this website. Ziehlneelsen technique and iodine which is added after the application of crystal violet in the gram staining method.
Misinterpretation of the gram stain has led to misdiagnosis or delayed. The gram stain, the most widely used staining procedure in bacteriology, is a complex and differential staining procedure. Certain bacteria are stained with aniline dyes, like ge. Gram staining uses crystal violet to stain cell walls, iodine as a mordant, and a fuchsin or safranin counterstain to mark all bacteria. Impregnation it is the deposition of salts of heavy metals on or around cells, tissue constituents etc. Christian gram in 1884 devised a technique that is the most helpful and most widely used staining technique in bacteriology. Kits are available commercially, or formulas for reagents are listed in appendix b. The gram stain, the most widely used staining procedure in bacteriology. In differential staining, specimen is subjected to a series of stains dyes in which different organisms or different parts of the cell are stained differently so that they can be distinguished from each other. Gram staining principles gram staining is used to determine gram status to classify bacteria broadly. Jan 24, 2020 gram staining is a common technique used to differentiate two large groups of bacteria based on their different cell wall constituents. In spite of his defective method, his name lives on jay hardy, cls, sm ascp hile working with infected lung tissue.
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