Answer to Question #164409 in Microbiology for Jamie Dimabuyu

Question

Why decolorizing is the most critical steps in gram stain?

Answer 1

The decolorizing is the most critical step in gram staining. We use decolorizing solution either alcohol or acetone and it is generally time bound. This is because if we decolorize for longer period of time that is if we over-decolorize the cells, we may get red color gram positive cells, and thus can be read as false gram negative bacteria. While if we under-decolorize then it may fail to remove the crystal violet iodine complex and thus can give purple color and thus can be read as false positive bacteria.

Hence, to avoid any false results by gram staining, the decolorizing step is consider as the most critical steps.