The reaction of ortho-phthalaldehyde (OPA) with amino acids and proteins was investigated as a possible mode of action. Bacterial pellets (obtained by centrifugation) changed colour after exposure to OPA. These colours were more intense at alkaline than acidic pH. Acidic and alkaline OPA reacted with primary amino acids to form coloured products. The reaction rate accelerated with increasing pH. OPA increased the optical density of bacterial cell suspensions (an indication of protein coagulation or microbial surface or other changes in the opacity of cell constituents). The inhibition of ethylenediaminetetraacetic acid- and sodium lauryl sulphate-induced lysis was not as great as for glutaraldehyde (GTA), possibly indicating less cross-linking of amines. Interactions with primary amino groups of the outer envelope or cell wall probably play a part in the action of OPA but the level of cross-linking associated with the outer membrane does not appear to be as extensive as that of GTA. The aromatic component might allow OPA to penetrate the outer layers of cells, thus helping to explain the very high activity of OPA against Gram-negative vegetative organisms even though the degree of cross-linking seems to be less than that seen with GTA. Thus, OPA reacts strongly with primary amines and stabilizes, to some extent, the outer membrane and cell walls of vegetative organisms and this probably accounts for part, but not necessarily all, of its lethal action.