Several monoclonal antibodies (McAbs) to human class II antigens are described; three of these recognize monomorphic epitopes, and two are polymorphic. The biochemical nature of the reactive epitopes was investigated by determining the binding ability of the McAbs to protease- or glycosidase-treated antigen preparations. Both of the polymorphic McAbs recognized protein-defined epitopes. However, for one of the antibodies F5C9, carbohydrate residues were also implicated. The extent of carbohydrate involvement varied with class II antigens of different DR specificity, suggesting that the proximity of the F5C9 epitope to carbohydrate side chains can vary. One of the three monomorphic McAbs recognized a protein-defined epitope, while the other two antibodies detected epitopes in which carbohydrates were involved. These results indicate that both protein and carbohydrate residues of the human class II antigens can influence the epitopes recognized by murine McAbs. This could explain some of the apparent complexities of the anti-human class II McAbs.