Full Answer
Why is heat used to stain fast bacilli?
Apr 14, 2020 · Heat fixing bacterial smears kills the bacterial cells so that they are fixed in place and ready for staining. Heat fixing also adheres the bacterial cells to the slide and allows the sample to take up the stain more easily. Fixation is a critical step in preparing slides and samples in microbiology, pathology and histology.
Why is it necessary to heat fix bacterial smears prior to staining?
Apr 27, 2018 · Why is Heat Used in Endospore Staining. The keratin covering of endospores resists staining. Therefore, the primary stain has to be forced into the endospore. The use of heat is to enhance the penetration of the primary stain into the endospore. The slide with malachite green can be heated up to 3-5 minutes.
Why is heat used to stain endospores?
Apr 04, 2020 · Heat is necessary when staining spores because the application of heat disrupts vegetative cells and causes green malachite to be rinsed from them, which then allows the counterstain into the cells. This is a required step in certain spores that are resistant to traditional methods of spore staining. According to Austin Community College, staining is a technique …
What is the heat staining technique?
This response may come a bit late but the purpose to heat kill any cell type / bacteria is to generate a positive control for dead-cell staining or on the other hand, negative control for any...
Why do you heat the stain bacteria?
The heat “melts” the waxy cell wall and permits the absorption of the dye by the cells. Then the slide is allowed to cool and a solution of acid and alcohol is added as a decolorizer. Cells that are “acid-fast” because of the mycolic acid in their cell wall resist decolorization and retain the primary stain.
Why do we heat fix bacteria before staining?
Heat fixing kills the bacteria in the smear, firmly adheres the smear to the slide, and allows the sample to more readily take up stains.
Why is heat fixation necessary for staining?
Heat fixing kills cells, and adheres them to the slide. Cells will be rinsed off the slides if they are not heat fixed properly.Mar 19, 2021
What does heat treatment do to bacteria?
The main objectives of heat treatment are enzyme inactivation to avoid decomposition reactions, and destruction of pathogens and spoilage microorganisms. Heating acts on at least one key enzyme of the bacterial metabolism; as a result microbial population reduction is a first-order kinetic reaction.
What are the two purpose of heat fixation?
Two purposes of heat fixation are: Kill the microbes. Living bacteria contain enzymes that can breakdown the structures of the bacteria. Heat...
Why is it necessary to stain bacteria before viewing in a microscope?
Why Stain Cells? The most basic reason that cells are stained is to enhance visualization of the cell or certain cellular components under a microscope. Cells may also be stained to highlight metabolic processes or to differentiate between live and dead cells in a sample.Feb 2, 2022
What is the purpose of the smear preparation?
The preparation of a smear is required for many laboratory procedures, including the Gram-stain. The purpose of making a smear is to fix the bacteria onto the slide and to prevent the sample from being lost during a staining procedure.
Why is heat fixing not necessary in negative staining technique?
Heat damages certain cellular features including bacterial glycocalyx (capsules and slime layers), therefore we do not heat fix when negative staining. Bacterial capsules are soluble in water, so we do not rinse with water in capsule staining.
What is heat treatment?
Heat treatment is the process of heating metal without letting it reach its molten, or melting, stage, and then cooling the metal in a controlled way to select desired mechanical properties.Jul 7, 2020
Which are goals of pasteurization?
The general objective of pasteurization is to extend product shelf-life by inactivating all non-spore-forming pathogenic bacteria and the majority of vegetative spoilage microorganisms, as well as inhibiting or stopping microbial and enzyme activity.
Are bacteria heat sensitive?
Bacterial spores are usually far more heat resistant than vegetative cells; thermophiles produce the most heat resistant spores while those of psychrotrophs and psychrophiles are most heat sensitive.
What is the counterstain used for?
After decolorization, the counterstain safranin is used to stain the background. Two pathogenic genera known as Bacillus and Clostridium produce metabolically inactive endospores to resist adverse environmental conditions. Since endospores cause a number of lethal diseases, the identification of them is quite important in clinical samples.
What is an endospore?
An endospore is a resistant structure produced by some pathogenic bacterial genera such as Bacillus and Clostridium in order to survive under unfavorable environmental conditions. These unfavorable conditions can be heat, desiccation, radiation or chemicals.
How long does it take for endospores to form?
It may take 8-10 hours to complete the process. Endospores may reside inside the bacterial cell or may exist as free spores. Endospore formation is shown in figure 1.
How to decolorize a microscope slide?
2. Then, gently heat the slide for 3-5 minutes until the dye starts to evaporate. 3. Allow the slide to cool and wash it with water for decolorization. 4. Finally, after counterstaining, rinse the slide. 5.
Why do we use staining in microscopy?
Staining is a valuable technique used in microscopy to enhance contrast in the microscopic image. Stains are used to highlight structures in clinical specimens, often when viewed with the aid of different microscopes. Stains have different affinities for different organisms and are used to differentiate types of organisms or to view specific parts ...
What is Gram stain?
The Gram stain is complex and differential staining technique that remains a useful test performed in microbiology laboratories. The staining procedure differentiates organisms of the domain bacteria according to the cell wall structure.
What is endospore production?
Introduction#N#Endospore production is an important characteristic of some bacteria (such as Bacillus and Clostridium species); this allows them to resist environmental conditions such as extreme heat, chemical exposure, etc.#N#The following methods below may be used for the demonstration of spores in Gram positive bacilli.
What is the purpose of auramine phenol stain?
1 Auramine-phenol stain – 1 (acid fast bacilli) This staining technique is used to demonstrate the presence of acid-fast bacilli (Mycobacterium species). These organisms have waxy envelopes that make them difficult to stain and decolorize. A fluorescent stain is used in this method.
What is the counterstain used for?
Neutral red, safranin, or carbol-fuchsin may be used as the counterstain. This technique has also been used for staining of certain fungi such as Candida and Cryptococcus which are observed as Gram-positive yeasts. Method.
Is phenol a carcinogen?
Disposable gloves must be worn when handling the reagents to avoid contact– Carbol fuchsin is carcinogenic while the acid-alcohol is corrosive. Phenol is a component of the carbol fuchsin reagent for the Ziehl-Neelsen and Kinyoun methods for acid-fast staining. Phenol is a dangerous chemical if not handled carefully.
What is Gram positive and Gram negative?
A culture containing Gram-positive and Gram-negative organisms may be used for quality control. Common errors in the Gram staining procedure. These are the errors that arise depending on the method and techniques used and which could result in a Gram-positive organism staining Gram negatively. They include;
What is the purpose of simple staining?
The purpose of simple staining is to elucidate the morphology and arrangement of bacterial cells. The most commonly used basic stains are methylene blue, crystal violet, and carbol fuchsin. Cultures: 24-hours nutrient agar slant culture of E.coli and Bacillus cereus , and a 24-hour nutrient broth culture of S. aureus.
Why is nuclear staining important?
Microbes are invisible to the naked eye and are difficult to see and identify, even when using a microscope. Staining microbes makes them easier to observe and reveals the presence of microscopic structures, because charged portions of the stain bind to specific macromolecules within the structures.
Why is it so difficult to visualize microorganisms in the living state?
Visualization of microorganisms in the living state is very difficult, not just because they are minute, but because they are transparent and almost colorless when suspended in an aqueous medium.
What are the two types of staining?
Stains are of 2 types: Acidic stains e.g., picric acid. Basic stains e.g., methylene blue. Types of staining techniques: Simple staining (use of a single stain) : This type of staining is used for visualization of morphological shape (cocci, bacilli, and spirilli) and arrangement (chains, clusters, pairs, and tetrads).
What is differential staining?
A simple stain displays the microorganisms, and a differential stain displays the chemical differences in cellular structures, including the cell wall and cell membrane, because the macromolecules within the structure bind to different components of the stain. An example of this differential staining is seen in staining used for blood smears.
What is a slide made of?
Slides can be made from direct clinical material (a wound, sputum, knee fluid, the throat, etc.), broth cultures, and solid media cultures. The first principle is that some fluid is needed to emulsify the material if it is dry; however, too much fluid may make the microbes hard to find.
Why do bacteria need bright field microscopy?
This is because of the lack of contrast coupled with the small size of the bacterial cell. The use of stains that react chemically with cell material will enhance the contrast between the cell and the background. A stain is a dye consisting of a colored ion (a chromophore) and a counter ion to balance the charge. Attachment of the chromophore part of the dye complex to a cellular component represents the staining reaction. Depending upon the dye, the chromophore can be either positively charged (cationic) and have an affinity for negative ions or negatively charged (anionic) with an affinity for positive ions. Bacteria carry a net negative charge at pH 7. Therefore, cationic dyes such as methylene blue, basic fuchsin, or crystal violet are useful for the direct staining of cells, whereas anionic stains, such as eosin and nigrosin, will not directly stain bacterial cells. However, negatively charged stains, are useful for revealing the outlines of bacterial cells; anionic dyes stain the background, leaving the bacterial cells clear and bright against a dark background.
How to give maximum light to stained slides?
For this you will need to give maximum light by opening the diaphragm, raising the condenser as much as possible, and adjusting the light source intensity.
