The mouse monoclonal antibodies AbA123, AbA127, AbK152, AbM68, and AbV1 were derived after immunization with cultured human tumor cells or melanocytes. Antibodies AbA123, AbA127, and AbK152 recognize human cell surface antigens expressed on most cultured human cells and show an identical pattern when tested on a panel of 47 human cell lines. They recognize at least two different epitopes on the same glycoprotein complex, designated A123/A127, which consists of 30,000-and 40,000-mol-wt glycopeptides. Antigens M68 and V1 are also expressed on most cultured human cell types but show distinct patterns of distribution on the cell line panel. The antigens defined by AbM68 and AbV1 have the characteristics of glycolipids. They are heat stable, and immunoprecipitation of metabolically labeled cell lysates did not yield any detectable components when analyzed by SDS-polyacrylamide gel electrophoresis. Serological typing of a panel of 23 independently derived mouse-human and Chinese hamster-human somatic cell hybrids showed unequivocally that the expression of cell surface antigens A123/A127, M68, and V1 segregates with human chromosome 12. The analysis of hybrids containing karyotypically defined deletions of chromosome 12 permitted the assignment of the loci determining the expression of antigens. A123/A127 and V1 to region 12cen-qter, and the locus determining the expression of antigen M68 to region 12cen-pter. These antigens can be distinguished from the cell surface molecules previously assigned to chromosome 12 and thus represent new assignments to this chromosome.