Bypassing hybridoma technology: HLA-C reactive human single-chain antibodyfragments (scFv) derived from a synthetic phage display library (HuCAL) and their potential to discriminate HLA class I specificities

The generation of discriminative, monospecific anti-HLA anti bodies used to be a difficult endeavor. Phage display technology, using single-chain anti body fragments (scFv) offers a powerful alternative obtaining target-specif ic, genetically stable reagents. Most of scFv obtained to date have been en riched by panning phage libraries to solid-phase coupled antigens, In the p resent study, HLA-C-specific scFv were isolated using a synthetic phage lib rary in combination with a Cw*0602 overexpressing cell line. ScFv from this procedure precipitated HLA-Cw*0602 heavy chains from whole cell lysates. F low cytometry analysis revealed that scFv stained HLA-Cw*0602-positive cell s, but not cells expressing HLA alleles Cw*0302 Cw*0802, A*0201, B*2705, or Gm1*01011, indicating the specificity of scFv. Similarly they showed an ab ility to discriminate Cw*0602-positive from Cw*0602-negative peripheral blo od lymphocytes (PBL). The rsults of our study demonstrate the feasibility t o genetically engineer single-chain HLA-class I-specific antibodies, by pha ge display technology. This approach might be a valuable tool to develop a broad range of novel monospecific antibodies against HLA-class I specificit ies.