CSMLS Practice Exam 2025 – All-in-One Guide to Mastering Medical Laboratory Science Certification

The counterstain used in Gram's stain is safranin. In the Gram staining process, after the initial application of the crystal violet stain, iodine is added, which forms a complex with the crystal violet, enhancing its retention in the gram-positive cells. This is followed by a decolorization step, usually with alcohol, which removes the crystal violet from gram-negative bacteria, rendering them colorless. At this stage, safranin is applied as the counterstain, which stains the decolorized gram-negative bacteria, allowing them to be visualized under the microscope. It provides contrast, enabling the distinction between gram-positive organisms, which retain the purple color from crystal violet, and gram-negative organisms, which take up the pink hue from safranin. This differentiation is fundamental in microbiology for identifying and categorizing bacterial species based on their cell wall characteristics.