gram staining

Meaning: Gram staining sorts bacteria into two groups by cell wall structure. Steps: crystal violet (primary stain), iodine (forms a large complex with the dye), alcohol or acetone (decolorizer), then safranin (red counterstain). Gram-positive bacteria have a thick peptidoglycan layer that traps the crystal violet–iodine complex, so they stay purple. Gram-negative bacteria have a thin peptidoglycan layer beneath an outer membrane containing lipopolysaccharide (LPS). The decolorizer disrupts the outer membrane and the dye washes out, so these cells take up safranin and look pink-red.

On the MCAT: A passage gives a stain result or a micrograph and asks you to infer wall structure, or to predict antibiotic response. Penicillin-type drugs block peptidoglycan cross-linking, and the outer membrane of gram-negative bacteria often limits drug entry. LPS in the outer membrane acts as an endotoxin. Stain color is often paired with shape (cocci, bacilli, spirilla) to classify a bacterium.

Common confusion: Gram-negative does not mean "no peptidoglycan." Both types have it; gram-negative cells just have a thinner layer plus the outer membrane. The decolorizing step is the critical one, so over-decolorizing can make gram-positive cells look gram-negative. Some bacteria, such as mycoplasmas, lack a cell wall and do not fit the usual pattern.