Scope of document the document covers the methods for the staining procedures commonly used in clinical microbiology laboratories for the identification of pathogens and dyes stains. When it is necessary to demonstrate whether or not a particular organism possesses a capsule, is flagellated, or forms endospores, special staining techniques must be used. Choose from 500 different sets of microbiology staining techniques flashcards on quizlet. Staining and streaking protocols for simple stain, gram stain, streak plate technique and culture maintenance. Microscopy using the microscope 22 stained preparations 23 making a smear 23 a simple stain 24 a differential stain. Introduction as bacteria consist of clear protoplasmic matter, differing but slightly in refractive index from the medium in which they are growing, it is difficult with the ordinary microscope, except when special methods of illumination are used, to set them in the unstained condition. Make a thin film of the material on a clean glass slide, using a sterile loop or swab for viscous specimens. Special stains differential stains basic techniques of. Gram stain is a very important differential staining technique used in the initial characterization and classification of bacteria in microbiology. Stains and dyes are frequently used in biological tissues for viewing, often with the aid of different microscopes. Staining, therefore, is of primary importance for the. The end result is a report that should assist the clinician in reaching a definitive diagnosis and a decision on antimicrobial therapy. This manual has been written for a righthanded person.
Gram staining is a differential staining technique that differentiates bacteria into two groups. Used to study small particles like viruses or macromolecules. Gram staining differentiates the bacteria into 2 groups. A simple stain will generally make all of the organisms in a sample appear to be the same color, even if the sample contains more than one type of organism. Hematoxylin and eosin staining technique functions to recognize different types of tissues and their morphological changes, especially in cancer diagnosis. Module context this module introduces microbiological laboratory techniques to be used for analysis of coliforms bacteria can be used as indicators of pollution. Sep 19, 2016 gram staining procedure animation microbiology principle, procedure, interpretation duration. Staining methods simple staining, negative staining, grams.
Histologic staining options available include, but are not limited to. A good report will likely necessitate a minimum of 2 pages doublespaced and is limited to a 3page maximum. By exploiting this phenomenon, an alternative gram staining technique has been developed. Fluorescencelabeled wheat germ agglutinin binds specifically to nacetylglucosamine in the outer peptidoglycan layer of grampositive bacteria.
Gram staining procedure animation microbiology principle, procedure, interpretation duration. In addition to staining procedures, biochemical tests used for differentiating bacteria. Basic techniques in biochemistry, microbiology and molecular biology pp 111116 cite as. Staining techniques in microbiology linkedin slideshare. A simple stain will generally make all of the organisms in a sample appear to be the same color, even if the sample contains more than one. Stains are salts composed of a positive ion cation and a negative ion anion. Under anaerobic conditions, grampositive staining occurred in all test organisms with quick fixing techniques with both absolute methanol and formalin. Diagnostic microbiology involves the study of specimens taken from patients suspected of having infections. To use and relate the gram stain to the study of bacterial cell morphology, and as an. 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. This stains the decolorized cells pink and is less noticeable in the cells that still contain the crystal violet dye. Types of staining techniques used in microbiology and their. This technique was introduced in 1884 by danish physician christian gram. In simple staining, a single dye is used to emphasize particular structures in the specimen.
Feb 03, 2015 staining techniques in microbiology 1. In microbiology the concept of staining is very important because it highlights the structures of microorganisms allowing them to be seen under a microscope simple and. Gram stain report 15 pts assignment each student will have the opportunity to submit one example of a gram stain with a brief written report describing the staining procedure and the stain itself. Alternate gram staining technique using a fluorescent lectin. These are of two types, namely direct and indirect staining. Staining techniques in microbiology archives learn. Gram staining helps to identify bacterial pathogens in specimens and cultures by their gram reaction grampositive and gramnegative and morphology coccirod. Neelsen in 1883 used ziehls carbolfuchsin and heat then decolorized with. Accurate gram staining of american type culture collection strains of these anaerobic bacteria is possible by implementing fixing and staining techniques within a gloveless anaerobic chamber. Some involve a single stain and just a few steps, while others use multiple stains.
Pdf correctincorrect ratios in staining techniques in. Microbiology module common staining technique microbiology 24 notes indeterminate groups as well. Choose from 500 different sets of staining techniques microbiology flashcards on quizlet. Microbiology module common staining technique microbiology 22 notes 2. Staining can be performed with basic dyes such as crystal violet or methylene blue, positively charged dyes that are attracted to the negatively charged materials of the microbial cytoplasm. To perform this staining, it requires the use of a single stain only. Special stains some bacteria have characteristic surface structures such as capsules or flagella and internal components e. Microscopy refers to the practice that involves the use of a microscope for the purposes of observing small scale structures that cannot be viewed using the naked eye and often cell staining is necessary as s tructures are difficult to discern due to insufficient contrast. Coloring microorganisms with a dye that emphasizes certain structures.
To study biological process like phagocytosis, molecular transport across plasma membrane. A detailed discussion of the different types of physical and chemical methods of sterilization is provided in chapter 19 physical and chemical methods of control. The gram stain is a differential stain commonly used in the microbiology laboratory that differentiates bacteria on the basis of their cell wall structure. Bacterial classification, structure and function introduction the purpose of this lecture is to introduce you to terminology used in microbiology. Diagnostic microbiology and laboratory methods pocket.
Comparison of two silver staining techniques for detecting. Staining is a supplementary method that gives divergence to the microscopic image for better vision under the microscope. There are many different stains and staining procedures used in microbiology. There are a variety of staining methods like simple, differential and special staining that are used in labs to examine the bacteria. A stain is a dye consisting of a colored ion a chromophore and a counter ion to balance the charge. Stains may be used to define biological tissues highlighting. The wide application of sterilization processes makes it necessary to impose strict control measures to validate the resu lts. Microbiology is a popular option for practical work in schools. The most widely used staining procedure in microbiology is the gram stain, discovered by the danish scientist and physician hans christian joachim gram in 1884. Hence, clinicians should be acquainted with the techniques of taking specimens, and understand the. Bhanu prakash usmle, fmge and neet pg 14,870 views. It is a technique that is widely used for the examination of cells, tissues and cellular components. Gram staining is the common, important, and most used differential staining techniques in microbiology, which was introduced by danish bacteriologist hans christian gram in 1884.
The peptidoglycan layer of gramnegative bacteria is covered by a membrane and is not labeled by the lectin. Techniques for controlling variability in gram staining of. Stains and dyes are frequently used in histology the study of tissue under the microscope and in the medical fields of histopathology, hematology, and cytopathology that focus on the study and diagnoses disease at a microscopic level. Microbiological laboratory techniques clinical gate. Some staining techniques involve the application of only one dye to the sample. Biochemical tests the sole purpose of biochemical test in microbiology is to differentiate between various classes of bacteria by studying their enzymatic activities. Cell staining in microscopy types, techniques, preparations. A gramstaining technique combining staining with two fluorescent stains, oregon greenconjugated wheat germ agglutinin wga and hexidium iodide hi followed by flowcytometric detection is described. Introduction to microbiology culture techniques youtube. This can be accomplished by physical methods such as heat, radiation, and filtration, or by chemical methods. Acidfast stain principle, procedure, interpretation and.
Flood slide with crystal or gentian violet 60 seconds. Procedure and uses august 1, 2014 acharya tankeshwar 0 bacteria are easier to detect in broth culture if the preparation is stained using toluidine blue 0. Testing sensitivity to antimicrobial substances 21 2. Gram stain is a very important differential staining techniques used in the initial characterization and classification of bacteria in microbiology. Gram staining gram staining is most widely used differential staining in microbiology. Reagents for histological techniques, haematology and. Two of the most important concerns are resolution and contrast.
Types of different staining techniques of microorganisms. It is one of the most important and widely used differential staining techniques in microbiology. Hence, clinicians should be acquainted with the techniques of taking specimens, and understand the principles and techniques. Some staining techniques, such as the gram and ziel neelsen stains, although. In the first step the smear is stained with basic dye crystal violet primary stain followed by treatment with iodine solution functioning as mordant. Uk standards for microbiology investigations issued by the standards unit, public health england. Staining microscopic specimens microbiology lumen learning. Staining simply means coloring of the micro organisms with the dye that emphasizes and elucidate different important structures of microorganisms including bacteria, virus, protozoa and etc.
Provide additional practice scenarios in staining techniques, with emphasis on tec hniques that require more than minute to apply a stain onto a smear e. Differential staining techniques viewing bacterial cells. Objectives to learn the technique of smear preparation. To learn the techniques of gram staining, nigrosin staining and koh test. Attachment of the chromo phore part of the dye complex to a cellular component represents the staining reaction. Microbiology module common staining technique microbiology 20 notes 2 common staining technique 2. Introduction observation of bacteria with conventional bright field microscopy yields relatively little useful information. Staining is a technique where specimens are stained with specific dyes to create contrast between the. So this method is also called ziehlneelsen staining techniques. Gram staining helps to identify bacterial pathogens in specimens and cultures by their gram reaction gram positive and gram negative and morphology coccirod. Neelsen in 1883 used ziehls carbolfuchsin and heat then decolorized with an acid alcohol, and counter stained with methylene blue. Staining techniques in microbiology a presentation by g.
Mcqs in microscopy and staining techniques part i category. It determines the cell shape, size and arrangement of the microorganism. The most commonly used instrument in the microbiology laboratory for the sterilization of media and glassware is the autoclave. This staining technique is used to demonstrate the presence of acid fast bacilli. Learn microbiology staining techniques with free interactive flashcards. Some staining techniques, such as the gram and ziel neelsen stains, although of great diagnostic value because of their differential staining properties for specific bacteria, reveal little.
Hematoxylin has a deep bluepurple color and stains nucleic acids by a complex, incompletely understood reaction. It is the differential staining techniques which was first developed by ziehl and later on modified by neelsen. Staining is a technique used to enhance contrast in samples, generally at the microscopic level. The microbiology society, in association with the microbiology in schools advisory committee misac. Staining procedures that use one dye to increase contrast between specimen and. Gramstaining is a differential staining technique that. This test differentiate the bacteria into gram positive and gram negative bacteria, which helps in the classification and differentiations of microorganisms. Learn staining techniques microbiology with free interactive flashcards. Methods of culturing microorganisms 3 five basic techniques 4 fig. Cover different classification schemes for grouping bacteria, especially the use of the gram stain 2. An optimized staining technique for the detection of gram. Gram staining in microbiology, fast staining in haematology, kit for staining reticulin fibres, pas staining and massons trichromic staining.
Smears were then blocked with 1 % bovine serum albumin bsa in trisbuffered saline tbs with 0. Then apart from all this some other staining techniques are there for identification of fungus like lactophenol cotton blue staining, and many other techniques. Jun 12, 2018 it is the differential staining techniques which was first developed by ziehl and later on modified by neelsen. Cell staining in microscopy types, techniques, preparations and procedures. The word gram is always spelled with a capital, referring to hans christian gram, the inventor of gram staining gram staining principles gram staining is used to determine gram status to classify bacteria broadly.
In microbiology the concept of staining is very important because it highlights the structures of microorganisms allowing them to be seen under a microscope. Staining of bacterial cells objective to learn the techniques of smear preparation, gram staining, nigrosin staining and correlating the results of gram staining with koh test. The classic silver staining method for detecting bacterial lipopolysaccharides lpss in polyacrylamide gels c. Make a thin film of the material on a clean glass slide, using a. Gram stain the previous lab introduced simple staining techniques that enable microbiologists to observe the morphological characteristics of bacteria.
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