Generation and selection of monoclonal antibodies, single-chain Fv and antibody fusion phage specific for human melanoma-associated antigens

Abstract

A panel of 13 murine monoclonal antibodies (mAbs) recognizing antigens on human melanoma cells but not on melanocytes was generated. Two mAbs (LHM3 and LHM5) stained sections of melanoma but not normal tissues. mAbs LHM2 and LHM8 stained only a minority of normal tissues. The mAbs differed further in their staining patterns on melanoma cell lines HMB2, DX3 and SK23 in FACS. The mAbs recognize antigens of 34, 38, 57, 94, 190-200 and > 200 kD. One mAb each bound to each of the antigens HLA DR (LHM4) and high molecular weight proteoglycan (LHM2). The high molecular weight proteoglycan-specific mAb was used to construct a single-chain Fv (scFv) antibody fragment and an antibody fusion phage in Escherichia coli. Both the scFv and the fusion phage were shown to bind specifically to melanoma cells. A method for the selection of melanoma cell-binding phages from phage libraries is described.