Staining in Microbiology [Microbiological Techniques [STM 312] Class one

                Staining Techniques

Why Staining?

Because microbial samples is usually transparent, it is necessary to stain micro-organisms before they are viewed with a light microscope. 

In some cases staining is a unnecessary for example when micro-organisms are very large or when motility is to be studied. In both cases, a drum of micro-organisms can be placed directly on a slide and observed, a preparation such as this is called wet mount.

     A wet mount can also be prepared by placing a drop of culture in a cover slip, and then inverting a cover slip over a slide that has a hollow in the center. This procedure is called the Hanging drop mount.

                 Staining Preparation.

   To prepare for staining, a small sample of micro-organisms is placed on a slide, and allowed to air dry, the smear is heat fixed by quickly passing it over a flame.

[Heat fixing: kills the organisms, makes them to adhere to the slide, and permit them to accept the stain].

Simple staining procedure...[picture gotten online].

The process of staining, involves an ion exchange reaction between the stain and the component to be stained e.g.

(i) bacteria cells are negatively charged, and to stain the bacteria cell, positively charged stains are used such that an ion exchange mechanism occurs.

           Composition of stain (dye)

      Every stain is composed of three components namely;

(1). Benzene ring

(2). chromophore

(3). Auxochrome

(1). Benzene ring: 

is the colorless part of the dye and it's the basic structural component of the dye.

(2). Chromophore:

is the functional group of a dye that gives the color to the stain.
Together, the benze ring and the chromophore is collectively knowm as the chromogen.

(3). Auxochrome: is the group that gives the ionic property to the stain.

        Types of stains based on chromogen.

      Based on the nature of the chromogen, there are three types of stains which are; 

1. The Acidic stain 

2. The Basic stain

3. The Neutral stain

1. The Acidic stain: are used to stain positively charged components of the background but they cannot stain bacteria cells, due to repulsion of the same charges. Examples include; 

i. eosin

ii. nigrosin and;

iii. india-ink.

2. The basic stain: the chromogen or the coloured part is positively charged and therefore can bound with negatively charged molecules such as bacteria cells. Basic dyes are most commonly used in bacteriology since the surface of bacteria cells are negatively charged e.g.

i. Crystal violet

ii. Methylene blue

iii. Safranin red

iv. Basic fuschin

3. The neutral dye: they are formed from the precipitation when acidic dye and basic dye are combined together, they are used to stain nucleic acids and the cytoplasm e.g.

i. Giemsa stain.

             Different staining Techniques.

Several types of staining Techniques include;

1. Simple stain

2. Differential stain

3. Capsule stain 

4. Acid-fast staining 

5. Spore staining 

6. Flagella staining. 

       

Thank you for staying tuned, continuation till next week... 

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