Negative Staining

In negative staining, actual specimen is left untouched and background is stained. This is the opposite of normal staining procedures in which actual specimens are stained. This technique is used to study cell morphological shape, size and arrangement, and also specific bacterial structures like the presence of diffuse capsules around many bacteria. In this staining procedure, bacteria are mixed with acidic dye like Indian ink or Nigrosin dye and then spread out on a slide as a thin film. It is then air dried and observed under microscope.

After giving up the hydrogen ion, the chromophore of the acidic dye becomes negatively charged. Since the bacterial surface is also negatively charged, it repels the dye and stains the background. As a result, the bacteria will show up as a clear spot against the dark background. This technique is also useful to stain those cells that are too sensitive to be heat fixed.