Introduction observation of bacteria with conventional bright field microscopy yields relatively little useful information. Bacteria have an affinity to basic dyes due to acidic nature of their protoplasm. 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. Some tests are used routinely for many groups of bacteria oxidase, nitrate reduction, amino acid degrading enzymes, fermentation or utilization of carbohydrates. The lab report assistant is simply a summary of the experiments questions, diagrams if needed, and data tables that should be addressed in a formal lab report. New staining technique for fungalinfected plant tissues.
To use and relate the gram stain to the study of bacterial cell morphology, and as an important step in the identification of a bacterial species. 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 technique is also referred to as acid fast staining. Hence staining techniques are used to produce colour contrast. The methylene blue stain is a simple technique, staining everything blue. Different bacteria stain differently to a common staining procedure. In microbiology the concept of staining is very important because it highlights the structures of microorganisms allowing them to be seen under a microscope. Gram stain is a very important differential staining techniques used in the initial characterization and classification of bacteria in microbiology. Most bacteria are identified and classified largely on the basis of their reactions in a series of biochemical tests.
Cover different classification schemes for grouping bacteria, especially the use of the gram stain 2. Method of staining used to distinguish and classify bacterial species into two large groups. Some staining techniques utilize these differences to stain the bacteria differently. Virology techniques introduction virology is a field within microbiology that encompasses the study of viruses and the diseases they cause. In negative staining technique, an acidic stain such as nigrosin, india ink, eosin or congo red is used in which the bacterial culture or the specimen is mixed well and then spread over the microscopic glass slide to form a thin smear. Grampositive bacteria gramnegative bacteria provides useful information for disease treatment. Gram stain developed in 1884 by hans gram, a danish microbiologist. Staining techniques in microbiology a presentation by g. In the past, conventional identi fi cation of fungi relied on the combination of morphological and physiological properties.
Gram stain protocols american society of microbiology. While some eucaryotes, such as protozoa, algae and yeast, can be seen at magnifications of 200x400x, most bacteria can only be seen with x magnification. Gram stain or gram staining, also called grams method, is a method of staining used to distinguish and classify bacterial species into two large groups. Classification of bacteria basic microbiology online. Routine methods of staining of bacteria involve dying and fixing smears procedures that kill them. 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. Already available was a staining method designed by robert. Staining is a technique used to enhance contrast in samples, generally at the microscopic level. Gram staining is a differential staining technique that differentiates bacteria into two groups. Crystal violet may also be used as a simple stain because it dyes the cell wall of any bacteria. This staining technique is used to demonstrate the presence of acid fast bacilli. Fixation of clinical materials to the surface of the microscope slide either by heating or by using methanol. In this exercise, bacterial endospores will be visualized by special staining techniques. 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.
Live bacteria do not show the structural detail under the light microscope due to lack of contrast. The previous lab introduced simple staining techniques that enable microbiologists to observe the morphological characteristics of bacteria. If this is your first attempt to access the mywgu portal, you need to create an account before you can sign in. It is almost always the first test performed for the identification of bacteria. Gram staining detects a fundamental difference in the cell wall composition of bacteria. Stains may be used to define biological tissues highlighting. Gram staining helps to identify bacterial pathogens in specimens and cultures by their gram reaction gram positive and gram negative and morphology coccirod. Negative staining technique principle, procedure, results. Gram staining is a common technique used to differentiate two large groups of bacteria based on their different cell wall constituents. Types of staining techniques used in microbiology and. Types of bacterial staining techniquespdf types of. Other bacteria lose the stain and take on the subsequent methylene blue stain blue. Make a thin film of the material on a clean glass slide, using a sterile loop or swab for viscous specimens. However, the use of these dyes by themselves only stains fungal structures andor plant cell walls and provides no clear distinction between the fungal and plant cell walls.
Objectives to learn the technique of smear preparation. The acid fast bacteria have a waxy substance mycolic acid on their cell wall that makes them impermeable to staining procedures. Before you can create an account, you must have filled out the wgu application and paid the application fee. A wet mount technique is used for staining bacterial flagella, and it is simple and useful when the number and arrangement of flagella are critical to the identification of species of motile bacteria. The name comes from the danish bacteriologist hans christian gram, who developed the technique. 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.
Capsule staining principle, reagents, procedure and result. To prevent the bacteria from washing away during the staining steps, the smear may be chemically or physically fixed to the surface of the slide. Gramnegative bacteria tend to exhibit lower fluorescence intensity than grampositive bacteria, because of their cell structure outer membrane impedes penetration of cfda. Capsules are fragile and can be diminished, desiccated, distorted, or destroyed by heating. In gram positive bacteria, the crystal violet dye iodine complex combines to form a larger molecule which precipitates. The gram stain is a differential method of staining used to assign bacteria to one of two groups grampositive and gramnegative based on the properties of their cell walls. Choose from 500 different sets of microbiology staining techniques flashcards on quizlet. Such staining methods are called differential staining methods, these include gram staining and acid fast staining. Classic gram staining techniques involve the following steps. The steps of the gram stain procedure are listed below and illustrated in table 1. Flagella stain principle, procedure and result interpretation. 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. The primary stain of this method is crystal violet, which. The procedure is named for the person who developed the technique, danish bacteriologist hans christian gram.
Histologic staining options available include, but are not limited to. Kits are available commercially, or formulas for reagents are listed in appendix b. In recent years, morphological studies, supplemented with staining techniques and biochemical methods, still play an. Bacteria and cells are visualised without enabling differentiation of bacteria according. View types of bacterial staining techniques pdf from biol 240l at university of waterloo. In 1884, hans christian gram, a danish doctor, developed a differential staining technique that is still the cornerstone of bacterial identification and taxonomic division. Staining techniques in microbiology linkedin slideshare. 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.
To be visible on a slide, organisms that stain by the gram method must be present in concentrations of. Make a thin film of the material on a clean glass slide, using a. The gram stain and differential staining journal of. By counterstaining with dyes like crystal violet or methylene blue, bacterial cell wall takes up the dye. The name comes from the danish bacteriologist hans christian gram, who developed the technique gram staining differentiates bacteria by the chemical and physical properties of their cell walls. 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. Other stain techniques seek to identify various bacterial structures of.
Distinguishes bacteria of two large and medically important groups. Introduction bacteria are too small to see without the aid of a microscope. Bacterial classification, structure and function introduction the purpose of this lecture is to introduce you to terminology used in microbiology. It was developed by danish microbiologist hans christian gram in 1884 as an effective method to distinguish between bacteria with different types of cell walls, and even today it remains one of the most frequently used staining techniques. In the laboratory, viruses have served as useful tools to better understand cellular mechanisms. A capsule in bacteria is the result of amorphous viscid secretion released by the bacteria. The gram stain procedure distinguishes between gram positive and gram negative groups by coloring these cells red or violet. Staining microscopic specimens microbiology lumen learning. When this secretion remains loose and undemarcated it is called a slime layer and when it is organized into a sharply defined structure it is called as capsule. The purpose of this lesson is to provide a general overview of laboratory techniques used in the. Capsules appear colourless with stained cells against dark background. The most useful staining procedure in medical microbiology. Gram stain the previous lab introduced simple staining techniques that enable microbiologists to observe the morphological characteristics of bacteria.
Some staining techniques involve the application of only one dye to the. To learn the techniques of gram staining, nigrosin staining and koh test. This test differentiate the bacteria into gram positive and gram negative bacteria, which helps in the classification and differentiations of microorganisms. Identifying bacteria using gram, acidfast and spore. The staining procedures require the use of a mordant so that the stain adheres in layers to the flagella, allowing visualization.
Types of different staining techniques of microorganisms. Learn microbiology staining techniques with free interactive flashcards. Bacteria vary in certain physiological and biological properties. Pdf staining techniques and biochemical methods for the. Other stain techniques seek to identify various bacterial structures of importance. A positive capsule stain requires a mordant that precipitates the capsule. Gram stain pure strain cultures to determine whether gram negative or gram positive. Grams iodine acts as a mordant helps to fix the primary dye to the cell.
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