A series of overlapping fragments spanning the entire coding sequence of the gH gene of murine cytomegalovirus (MCMV) were expressed in Escherichia coil as fusion proteins with glutathione S-transferase (GST) using the pGEX expression system. A region of antibody-binding was mapped to the NH2-terminus of glycoprotein H (gH) between amino acid residues 26 and 90 on the basis of the reactivity of GST-gH fusion proteins with polyclonal antibodies to MCMV in Western blot analysis. Antibodies to gH were generated in mice immunized with the GST-gH fusion protein SK and shown to react with an 87-kDa polypeptide present in virion particles which was conserved in MCMV isolates obtained from diverse locations. They also recognized the gH protein in MCMV-infected cells, as well as gH expressed in Chinese Hamster Ovary cells. The antibodies to gH had a significant ELISA titer but no neutralizing activity in vitro. The antibody response to the GST-gH fusion protein did not modify the level of MCMV replication in the spleens of mice. (C) 1994 Academic Press, Inc.