In the non-specific Neisser procedure, methylene blue, crystal violet and chrysoidine are used for the detection of metachromatic granules, or so-called Babes-Ernst polar bodies, especially in diphtheria bacteria.

With a defined pH value, methylene blue and crystal violet will be bound in the polar bodies or structures (Volutin bodies), but not bound in the rest of the bacterial cell. The polar bodies become visible as dark dots.

In the counter stain procedure, the body of the bacterium is stained by chrysoidine but this is only partly absorbed by the polar bodies.

Neisser solution can be carried out with various specimens such as air-dried, heat-fixed smears of bacteriological material, e.g. sputum, FNAB, lavages, imprints, body fluids, pus, exudates, liquid and solid cultures.

The 3 Neisser solutions from Merck are ready-to-use and Neisser solution Ia and Ib are mixed immediately prior to use in a ratio 1:1, Neisser solution II is used for the counter stain.

For the modification acc. to Gin, there is a preparation of Lugol's solution, mixed with lactic acid. Details are described on the insert sheet that is available on-line at
microscopy.merck.de

This established staining protocol gives good and reproducible results:
Stain for 1 min in methylene blue-crystal violet solution
Wash very briefly - or not at all - with water
Counter stain immediately with Neisser's solution II for 10 sec
Wash briefly with water and dry between sheets of blotting paper

Result
Polar bodies dark brown to black
Bacterium body brownish

Ready-to use stain solutions can visibly improve the quality of stains used routinely.

Available reagents

For further information about the wide range of products available from Merck KGaA for bacteria detection by staining, please contact Merck KGaA (see details above) or EMD Chemicals in the USA or click the link below.