Binding characteristics of CA 15-3 assay antibodies DF3 and 115D8 to MUC1 mucin

The CA 15-3(R) assay measures serum MUC1 mucin. This assay has shown clinic al utility in the management of patients with metastatic breast cancer. Man y commercial versions of CA 15-3 assays are available, and discrepant resul ts are found among these assays for the same patient samples. Although the DF3 and 115D8 monoclonal antibodies (MAbs) were first described in the 1980 s, a detailed understanding of the binding of these MAbs with MUC1 mucin is lacking. Therefore, we compared the binding, pH dependence, and epitope sp ecificity for both DF3 and 115D8. MUC1 mucin from the serum of cancer patie nts contains fewer sialyl terminals than mucin from healthy individuals. We partially removed sialic acid residues from MUC1 mucin with neuraminidase to mimic the less sialylated forms observed in cancer patient sera. Our res ults showed that DF3 and 115D8 recognize distinct epitopes and bind to MUC1 mucin with low apparent affinity. Additionally, both MAbs demonstrated red uced binding to desialylated MUC1 mucin. We also fractionated conjugates of DF3-alkaline phosphatase (DF3-ALP) by gel filtration and found that the bi nding to untreated mucin was similar across the molecular size gradient. Ho wever, significant differences in antibody binding to desialylated mucin we re seen for larger sized species of antibody conjugates compared with small er sized conjugates. These results suggest that conjugation of the DF3 anti body with ALP may alter recognition of differently sialylated forms of MUC1 mucin. This understanding may have utility in the standardization of CA 15 -3 assays.