Staining solutions

Accentuators & mordants :

In order to increase the intensity of staining or to stain at all it may be necessary to add an accentuator or intensifier to a staining reagent. At times heat may be used successfully.chemical accentuators are generally acidic or basic in nature and include such substances as acetic acid, oxalic acid, phenol, potassium hydroxide, soaps , aniline and formaldehyde. Some of these materials may serve in a dual role as fixatives and as accentuators.

Most of the intensifiers act by affecting the dissociation of ampholytes present in the cell. Accentuators are to be distinguished from mordants by the fact that they do not act by combining with the dye. Mordants on the other hand are chemical substances which have the power of making dyes stain materials otherwise unstainable or increasing their affinity or selectivity for particular structures. Mordants have a strong affinity for both substrate and dye and thus in a literal sense they can anchor a dye to a substance.

The union of mordants and dye can be demonstrated in vitro. Such reaction products are less soluble than the reactants, and resulting precipitates are termed lakes. In the presence of mordants, dyes may be monogenetic showing one,color, or polygenetic giving various color with different mordants.

Sometimes mordants is applied before the stain solution while in some other cases it may be applied along with stain. Tannic acid, salts of aluminium, tin, zinc, copper and chromium.

Staining the living cell:

Usually cells are stained after killing them. Since many dyes are highly toxic their application in sufficient concentration commonly results in death. If the application of the stain kills the cell, this type of staining applied to previously unfixed living cells is called supravital staining.

Distortion of normal structure by fixatives may often be evaluated by comparison against supravital staining. Janus green is one of the most useful supravital stains. Supravital staining has been used as in the proca-kayser method to distinguish dead from living bacteria. Methods of a similar kind have also been suggested for telling dead from living bacterial endospores.

Organisms may also be stained vitally, this vital staining, also known as intravital staining, being defined as the non toxic coloration of cell.