Before staining and observing a microbe under a microscope, a smear must be prepared. The goal of smear preparation is to place an appropriate concentration of cells on a slide and then cement them there so that they do not wash off during the subsequent staining procedure.
What are two important qualities of a good bacterial smear?
1. Cells that adhere to the slide and does not wash off during staining and washing procedures . 2. No shrinkage of cells.
Why is it important to prepare a good bacterial smear?
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 . A smear can be prepared from a solid or broth medium.
Why is it essential to prepare a bacterial smear for Gram staining?
Please note: It is very important to prevent preparing thick, dense smears which contain an excess of the bacterial sample . A very thick smear diminishes the amount of light that can pass through, thus making it difficult to visualize the morphology of single cells.
What is the purpose of fixing the bacterial smear?
The purpose of making a smear is to fix the bacteria onto the slide . Fixing the bacteria will preserve the morphology of the cells long-term. Also, fixation assists the cells in adhering to the slide, so that the cells do not fall off the slide during the staining procedure.
How do you properly prepare a bacterial smear?
- Label a clean slide with the initials of the organism.
- Using a loop, place one to two loops of water on center of slide.
- With a sterile loop, touch the entire loop to the culture and emulsify the cells in water on Slide.
- Allow all slides to air-dry completely.
Why is it important that you don’t put too much water to prepare the smear?
The preparation of a bacterial smear can be difficult to learn. ... When mixing bacteria with water, if you over mix it can result in inaccurate cell patterns. 3. When heat-fixing, if you hold the slide over the flame too long, you will incinerate the cells.
What are ideal qualities of a bacterial smear?
An ideal bacterial smear should be thin, semi-transparent, whitish layer or film, circular with diameter 1 cm. (approximately) free from dirt , dust or any contamination.
What are the two types of smear preparation?
Four different types of smear preparation methods (conventional method, blood film method, drop and rest method, and water-wash method) were carried out according to the standard reference as described below. A drop of sample was placed onto a clean slide and then spread to make a smear of 1 cm diameter.
What are some consequences of Overstaining a bacterial smear?
What are some consequences of leaving a stain on a bacterial smear too long (over-staining)? Consequences of over-staining are that the cell wall may be broken up or completely destroyed which would result in a loss of morphological characteristics of the bacterial cell .
What are the types of stains?
- Oil Stain. Oil stains are the most widely available and the type of stain most people think of when they think of stain. ...
- Varnish Stain. Varnish stains resemble oil stains in every way but one. ...
- Gel Stain. ...
- Lacquer Stain. ...
- Water-Soluble Dye Stain. ...
- Metal-Complex (Metalized) Dye Stain.
What is the most commonly used method in staining?
The Schaeffer-Fulton method (the most commonly used endospore-staining technique) uses heat to push the primary stain (malachite green) into the endospore. Washing with water decolorizes the cell, but the endospore retains the green stain. The cell is then counterstained pink with safranin.
What color is gram negative bacteria?
Gram negative organisms are Red . Hint; Keep your P’s together; Purple is Positive. Gram stains are never pink they are red or purple so you don’t destroy the rule; keep your P’s together. In microbiology bacteria have been grouped based on their shape and Gram stain reaction.
What happens if you overheat a bacterial smear?
If the smear is overheated during heat fixing, the cell walls will rupture . Concentration and freshness of reagents may affect the quality of the stain. Washing and drying of the smear between steps should be consistent. Excess water left on the slide will dilute reagents, particularly Gram’s iodine.
What is the importance of a properly stained smear?
A properly prepared smear accomplishes two things. It causes bacteria to adhere to a slide so that they can be stained and observed . It also kills them, rendering pathogenic bacteria safe to handle. An objective in preparing smears is to learn to recognize the correct density of bacteria to place on the slide.
Why is it important to limit the number of cells when making a bacterial smear?
why is it important to limit the quantity of cells used to prepare a smear? large amounts of cells in a smear can cause staining artifacts because stain is not washed away by destaining agents or water .
