Negative staining methode

NEGATIVE STAINING ::

principle: 

Negative staining use acidic dye like nigrosine, congo red. Acidic dyes have negatively charged chromogen which will not bind (react) with bacterial cells because of similar acidic chargr on the surface of bacterial cells . The stain does not stain the bacteria because of ionic repulsion. 

1. Natural shape and size of the cell is not disorted because the heat fixation is not required during the staining.

1. It is possible to observe those bacteria which are difficult to stain , such as spirochetes

1. requirements:

• Young culture of E. coli
• Nigrosin staining solution

PROCEDURE :

1.- place a small drop of nigrosine at the end of the slide. for culture on solid media, add a loopful of distilled water and emulsify a small amount of the culture in the Nigrosine-water drop. for broth cultures, mix full of the culture into the drop of Nigro Sine. do not spread the drop or let dry.

2. Using the end edge of another slide, spread the Drop out to produce a Samear varying from opaque black to gray. the angle of spreading slide will determine the thickness of the smear.

3. Let the smear air dry and examine ( do not  fix the slide)

RESULT:

● organism are observed as transparent objects against the dark background .

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