The Gram Stain and Differential Staining

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Abstract
Methyl alcohol solutions containing different amounts of I when used as decolorizing agents in the Gram test show Gram-positives to vary among themselves in degree of positivity and in different parts of the cell body. Sufficient quantities of I in ethyl alcohol prevented the decolorization of negatives. Without previous I treatment, small quantities of I in ethyl alcohol produced partial decolorization in Gram-positives, resulting in striking patterns. A difference between positives and negatives was observed on treatment with alkali, the latter being converted into slimy masses. While alkali treatment does not interfere with staining, changes produced in the interior by this treatment can be made visible by methylene blue and I and also by dye pairs such as crystal violet and safranin, using the latter to displace the former. In general, the displacement method with dye pairs was found to possess great possibilities for differentiating organisms.

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