The monoclonal antibodies B1.23.2 and B9.12 were developed to characterize human class I major histocompatibility gene products. They detect two different epitopes and define on a given lymphoblastoid B cell line two subsets of beta 2 microglobulin-complexed HLA class I molecules. One subset reacts with B9.12 and B1.23.2 and the other one expresses only the B9.12 epitope. Binding assays were performed on C3H mouse L cells which had been transformed with various single HLA-class I genes and the two detected molecular subsets were shown to be encoded by different genetic loci. Unlike B1.23.2, B9.12 detects all the beta 2m-complexed molecules expressed in human B cell lines and recognizes an epitope different from the one defined by W6/32. In contrast to the vast majority of the other reported anti-class I monoclonal antibodies (including B9.12), B1.23.2 recognizes an epitope expressed on both beta 2m-associated and -free HLA class I heavy chains.