title: Bypassing hybridoma technology: HLA-C reactive human single-chain antibody fragments (scFv) derived from a synthetic phage display library (HuCAL) and their potential to discriminate HLA class I specificities.
abstract: The generation of discriminative, monospecific anti-HLA antibodies used to be a difficult endeavor. Phage display technology, using single-chain antibody fragments (scFv) offers a powerful alternative obtaining target-specific, genetically stable reagents. Most of scFv obtained to date have been enriched by panning phage libraries to solid-phase coupled antigens. In the present study, HLA-C-specific scFv were isolated using a synthetic phage library in combination with a Cw*0602 overexpressing cell line. ScFv from this procedure precipitated HLA-Cw*0602 heavy chains from whole cell lysates. Flow cytometry analysis revealed that scFv stained HLA-Cw*0602-positive cells, 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 ability to discriminate Cw*0602-positive from Cw*0602-negative peripheral blood lymphocytes (PBL). The results of our study demonstrate the feasibility to genetically engineer single-chain HLA-class I-specific antibodies, by phage display technology. This approach might be a valuable tool to develop a broad range of novel monospecific antibodies against HLA-class I specificities.
authors.affiliation: Institut für Immunologie, Klinikum der Universität Kiel, Germany. marget@immunologie.uni-kiel.de
authors.forename: M
authors.initials: M
authors.lastname: Marget
authors.forename: B B
authors.initials: BB
authors.lastname: Sharma
authors.forename: M
authors.initials: M
authors.lastname: Tesar
authors.forename: T
authors.initials: T
authors.lastname: Kretzschmar
authors.forename: S
authors.initials: S
authors.lastname: Jenisch
authors.forename: E
authors.initials: E
authors.lastname: Westphal
authors.forename: P
authors.initials: P
authors.lastname: Davarnia
authors.forename: E
authors.initials: E
authors.lastname: Weiss
authors.forename: M
authors.initials: M
authors.lastname: Ulbrecht
authors.forename: D
authors.initials: D
authors.lastname: Kabelitz
authors.forename: M
authors.initials: M
authors.lastname: Krönke
mesh_terms: D000483:Alleles; D000918:Antibody Specificity; D002460:Cell Line; D005434:Flow Cytometry; D015236:HLA-C Antigens / Q000032:analysis* / Q000235:genetics / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008214:Lymphocytes / Q000737:chemistry; D019151:Peptide Library; D011233:Precipitin Tests; D014162:Transfection
chemical_list: D015236:HLA-C Antigens; D015395:Histocompatibility Antigens Class I; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library