Negative Stains

Negative Stains

Negative Stains

Negative staining is a technique by which bacterial cells are not stained, but are made visible against dark background.
Acidic dyes like eosin and nigrosin are employed for this method. Though, this staining technique is not very popular, it has an advantage over the direct or positive staining methods for the study of morphology of cells. This is because of the fact that the cells do not receive vigorous physical or chemical treatments.

The colouring power of acidic dye e.g. eosin in sodium eosinate is having negative charge, therefore, it does not combine with the negatively charged bacterial cell surface. On the other hand, it forms a deposit around the cell, resulting into appearance of bacterial cell colourless against dark background as shown in Figure.

For the staining, take a loopful of bacterial suspension on a slide and a drop of 10% nigrosin (G.T. Gurr) or 2 % congo red solution. Mix thoroughly with the help of sterile wire loop. Spread this mixture as a thin film across the slide using the edge of another slide. Allow it to air dry and examine the smear under oil-immersion objective.

Too thick layering of staining solution will not allow lights to pass through the object and dye will crack on drying. Too thin film will not give good contrast.

Most bacterial cells will be seen colourless against the dark back ground as shown in Figure. Some bacilli, such as those of coliform and haemophilic group, will show in their central portion a slightly dark patch some what resembling a nucleus.

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