Eleven different monoclonal antibodies specific for H-2K- and H-2D-encoded Class I molecules have been screened to determine Class I epitopes dependent on both carbohydrate and protein structures. Monoclonal antibodies have been identified which bind to carbohydrate-defined antigens encoded by both the H-2K and H-2D gene regions. Sensitivity to glycosidases versus pronase has been used to classify antigens both expressed as cell surface molecules and when prepared as detergent solubilized antigen. Several simple sugars have also been found to act as inhibitors of antibodies which bind to carbohydrate-defined sites. The genetic control of carbohydrate antigen expression by H-2K- and H-2D-linked genes has been verified since a specific antibody does not bind to H-2Kb or H-2Db molecules encoded by several mutant strains of mice containing single amino acid substitutions in their protein product. All of these data are consistent with Class I antigenic structures being encoded in carbohydrate as well as protein moieties.