In Gram staining, Gram's iodine serves as a mordant — it forms an insoluble complex with crystal violet (CV-I complex) within the bacterial cell, intensifying and fixing the stain. This CV-I complex is larger than crystal violet alone and is trapped within the thick peptidoglycan wall of Gram-positive bacteria after decolourisation, causing them to retain the purple colour. In Gram-negative bacteria, the CV-I complex is washed out by alcohol/acetone because their thin peptidoglycan cannot retain it.