What is the function of a counterstain in the Gram staining procedure?

The function of a counterstain in the Gram staining procedure is to stain the decolorized cells. During the Gram stain, crystal violet is used as the primary stain, which gives all the cells a violet color. After applying iodine, which binds to the crystal violet, a decolorization step follows where ethanol or acetone removes the crystal violet from certain cells, particularly Gram-negative bacteria. These decolorized cells require a subsequent staining step to make them visible under a microscope. The counterstain, usually safranin, provides this visibility by staining the cells that have lost their primary dye, thus allowing for differentiation between Gram-positive cells (which retain the violet color) and Gram-negative cells (which take on the color of the counterstain). This distinction is crucial for understanding bacterial classification and characteristics.