GENERATION OF MURINE MONOCLONAL ANTIHUMAN MILK-FAT GLOBULE-MEMBRANE ANTIBODIES USING IMMUNOPRECIPITATION AND BIACORE(TM) ANALYSES

A selection of monoclonal antibodies was developed against deoxycholin e-solubilized human milk fat globule membranes (HMFG). The antibodies were selected for their ability to immunoprecipitate I-125-labeled HMF G and then further analyzed by surface plasmon resonance on the BIAcor e(TM) for their reactivity with HMFG and with a fusion protein contain ing a 4-mer of the muc-1 tandem repeat. Both the HMFG and the fusion p rotein proved to be robust surfaces for the analysis of crude supernat ants. The BIAcore evaluation was useful in identifying true positives. BIAcore analyses of purified antibody preparations were used to deter mine binding characteristics such as affinity and intensity. The latte r proved useful in selecting a panel for evaluation by immunohistochem istry for breast tissue reactivity. Four of 6 antibodies appeared to r eact more intensely with tumor compared with normal breast tissues. On e of those antibodies reacted with the fusion protein 4-mer of the muc -1 tandem repeat.