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.
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.
Marget
M
M
Institut für Immunologie, Klinikum der Universität Kiel, Germany. marget@immunologie.uni-kiel.de
Sharma
B B
BB
Tesar
M
M
Kretzschmar
T
T
Jenisch
S
S
Westphal
E
E
Davarnia
P
P
Weiss
E
E
Ulbrecht
M
M
Kabelitz
D
D
Krönke
M
M
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
D015236:HLA-C Antigens; D015395:Histocompatibility Antigens Class I; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library