Production and characterization of a monoclonal antibody against the seed lectin of the Dolichos biflorus plant.

Spleen cells from mice immunized with the Dolichos biflorus seed lectin were fused with cells from the mouse myeloma Sp2/O-Ag14 cell line to form hybridomas. Those hybridomas producing antibodies against the seed lectin were cloned at least four times and the monoclonal antibodies from clone C11/64-56.28 were characterized and found to be specific for Subunit I of the lectin; they do not react with the structurally similar Subunit II. In previous studies, we have shown that although these two subunits appear to differ only at their COOH-terminal ends, only Subunit I has carbohydrate binding activity. Using a solid phase enzyme immunoassay, the antigenic determinant fr the monoclonal antibody was found to be located on the COOH-terminal cyanogen bromide fragment of this subunit. The monoclonal antibody inhibits the ability of the lectin to agglutinate erythrocytes and N-acetyl-D-galactosamine, the specific hapten for the lectin, inhibits the ability of the antibody to combine with the lectin. These results suggest that the monoclonal antibody recognizes a determinant that is located either at or near the active site of the lectin or that is conformationally interdependent with the active site.