AAT Bioquest

What is the role of alcohol in the Gram stain procedure?

Posted March 22, 2023


Alcohol is responsible for penetrating the cell walls of G- bacteria in order to dissolve the crystal violet-iodine complex. The dissolved peptidoglycan layer is thin and unable to retain the crystal violet-iodine complex and turns colorless. In contrast, G+ bacteria has a thicker peptidoglycan layer that is impenetrable to decolorizers like alcohol. Because of this, the crystal violet-iodine complex is retained in the cell walls of G+ bacteria and the cells will appear purple. In G+ bacteria, Alcohol dehydrates the peptidoglycan layer, shrinks it, and then tightens it. The crystal violet-iodine complex is unable to enter the tightened peptidoglycan layer and therefore stays permanently in G+ bacterial cells.    

Additional resources

Studies in Gram staining

Gram Staining

MycoLight™ Rapid Fluorescence Gram-Positive Bacteria Staining Kit