Which statement about Gram stain mordant is true?

• A. The mordant is applied after the primary stain
• B. The mordant is applied before the primary stain
• C. The mordant is the decolorizer
• D. The mordant is the counterstain

Answer: (A)

The main concept is how the mordant works to fix the stain. In Gram staining, the mordant (iodine) is applied after the primary stain (crystal violet) to form a crystal violet–iodine complex inside the cell wall. This complex is larger and less soluble, so it gets trapped in Gram-positive cell walls that have a thick peptidoglycan layer. When the decolorizer (alcohol) is applied, these Gram-positive cells stay purple because the complex remains bound, while Gram-negative cells, with thinner peptidoglycan and an outer membrane that’s disrupted by the alcohol, lose the dye. The final counterstain then colors the decolorized Gram-negative cells. The mordant is not the decolorizer and not the counterstain, and it is applied after the primary stain, not before it.