During a Gram stain, what is the effect of applying alcohol on Gram-negative cells?

During a Gram stain, the application of alcohol serves as a decolorizing agent. In Gram-negative cells, which have a thinner peptidoglycan layer and an outer membrane, the alcohol disrupts the outer membrane and allows the crystal violet stain to leach out. As a result, the Gram-negative cells become colorless after the alcohol is applied. This is a crucial step in differentiating between Gram-positive and Gram-negative bacteria, as the latter will subsequently take up the counterstain (safranin), resulting in a red appearance. Understanding this process is essential for recognizing how bacterial cell wall structure influences staining properties in microbiology.