IDENTIFICATION AND CHARACTERIZATION OF MONOCLONAL-ANTIBODIES THAT PARTIALLY BLOCK HUMAN NATURAL ANTIBODY-BINDING TO PIG ENDOTHELIAL-CELL XENOANTIGENS

The shortage of human donors for clinical transplantation has led to a serious consideration of the use of non-human species as organ donors . The major barrier to the clinical use of xenografts from species suc h as the pig in human transplantation has been the aggressive nature o f the immune-mediated rejection of the graft. We have recently identif ied the molecular weights of several endothelial cell surface proteins that may be targets of human antibody-mediated responses to pig aorti c endothelial cells (PAEC). In this series of experiments, we produced a panel of rat monoclonal antibodies (Mabs) to PAEC in an effort to i dentify Mabs that detect pig xenoantigens. Mabs were selected based on flow cytometric binding to PAEC, pig platelets, and various pig cell lines, including a pig kidney cell line (LLC-PK1) reported to react wi th human natural antibodies (HNA). Eleven of the eighty-three antibodi es produced were cytotoxic for PAEC. Six of the cytotoxic clones recog nized a 44 kDa protein and two of the clones recognized a 115 kDa prot ein expressed on the surface of PAEC. Since PAEC target antigens recog nized by human natural antibodies include both 115 and 44 kDa antigens , these Mab clones were selected for further study. Several distinct p atterns of tissue reactivity were demonstrated within this group of an tibodies by immunohistochemical analysis; however all monoclonal antib odies were highly reactive with endothelial cells in all tissues exami ned. Two monoclonal antibodies recognizing antigens that are highly ex pressed on pig endothelial cells (92-98%) and pig platelets (74-92%), but moderately expressed on pig splenocytes (33-38%), were capable of reproducibly blocking 48-53% of human IgM binding to pig endothelial c ells when analyzed with flow cytometry. This data suggests that these Mabs may recognize epitopes of potential significance in the human-to- pig xenograft reaction.