Gram Staining Procedure

Gram stain is a method of differential staining. Gram stain was discovered and introduced by Christian Gram in 1884. This divides bacteria into two groups, gram positive and gram negative, depending on the presence of peptidoglycan in their cell wall. Bacteria lacking peptidoglycan termed as gram negatives and gram positives are the bacteria who contain peptidoglycan. Peptidoglycan can absorb dyes hence the presence of peptidoglycan gives color. 

Gram staining method has two particular dyes. Primary dye is the ammonium oxalate crystal violet which stains all cells with purple. Mordant is a substance that strengthens the union between the dye and the substrate or fixes coloring material. Gram’s iodine solution used as the mordant. Decolorizing agent 95% alcohol removes the primary stain from gram-negative cells and gram positives retain the primary dye. Safranin a secondary dye is used to stain the colorless gram-negative cells. Hence gram-positive cells appear in purple whereas gram-negative cells show a color of pink to red. This differential behavior has been explained by; the cell wall composition of gram negatives and positives and differential permeability to alcohol insoluble dye, iodine complex.

Gram Reaction 

This method is to distinguish gram negatives and gram-positive bacteria. This comparison depends on the presence of peptidoglycan in the cell wall.

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