How does peptidoglycan affect gram staining?

A popular method for separating two sizable groups of bacteria according to the components of their respective cell walls is gram staining.

By coloring these cells red or violet, the Gram stain method helps to distinguish between Gram positive and Gram negative groups.

The reason gram positive bacteria stain violet is because their cell walls contain a thick layer of peptidoglycan, which keeps the crystal violet stain on the bacteria's surface.

On the other hand, the reason why Gram negative bacteria stain red is because their peptidoglycan wall is thinner and does not hold on to the crystal violet color during the decolorizing process.

Even if the lab technician believes she already knows the outcome, all steps are completed.

Three steps are involved in gram staining: decolorization, staining with a water-soluble dye called crystal violet, and counterstaining, which is typically done with safanin (light red).

Gram positive bacteria (with a thicker peptidoglycan layer) retain crystal violet stain during the decolorization process, while Gram negative bacteria lose the crystal violet stain and are instead stained by safranin in the final staining process. This is because the peptidoglycan layers in the cell membranes of Gram positive and Gram negative bacteria differ in thickness.

Peptidoglycan in bacterial cell walls retains crystal violet dye, influencing the retention or loss of color during the gram staining process. Gram-positive bacteria retain the dye, appearing violet, while gram-negative bacteria lose the dye and take up the counterstain, appearing red.