Quantification and identification of soluble HLA-G isoforms.
In order to clarify the diagnostic relevance of soluble human leukocyte antigen-G (sHLA-G) molecules, reliable methods for the measurement of sHLA-G in various body fluids are of interest. Therefore, the aims of the 'Wet-Workshop for Quantification of Soluble HLA-G' held in Essen, Germany (at the Institute of Immunology, 18-20 October 2004) were to select and to validate HLA-G-specific enzyme-linked immunosorbent assay (ELISA) formats and purified standard HLA-G proteins, which can be easily generated and used as consensual references. We chose two ELISA formats, one for the simultaneous determination of shed HLA-G1 + sHLA-G5 (sHLA-G1 + G5) and one for the exclusive detection of HLA-G5 molecules. The first ELISA uses the antibody pair monoclonal antibody (mAb) MEM-G/9 + anti-beta2-microglobulin (beta2m), whereas the latter uses mAbs 5A6G7 + W6/32. Purified and well-defined HLA-G5 protein derived from insect SF9 cells transfected with HLA-G5 + human beta2m served as standard reagent. Twenty-five members of 13 international laboratories participated in the 3-day Wet-Workshop. The workshop demonstrated that the HLA-G5 protein was equally detected by both ELISA formats allowing direct comparison of quantitative results obtained by these two ELISA formats, and that sHLA-G1 + G5 and HLA-G5 molecules, respectively, were specifically and reproducibly quantified by the two ELISA formats. The comparison of the two ELISA results obtained allows the conclusion that sHLA-G1 and HLA-G5 molecules can exist in the blood of healthy donors. Moreover, there was evidence for a novel soluble HLA-G structure recognized by the mAbs 5A6G7 + W6/32 antibody combination but not by the one of mAb MEM-G/9 + anti-beta2m.
Rebmann
V
V
Institute of Immunology, University Hospital of Essen, Essen, Germany.
LeMaoult
J
J
Rouas-Freiss
N
N
Carosella
E D
ED
Grosse-Wilde
H
H
D000653:Amniotic Fluid / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006680:HLA Antigens / Q000097:blood* / Q000276:immunology; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I / Q000097:blood* / Q000276:immunology; D006801:Humans; D010051:Ovarian Neoplasms / Q000097:blood / Q000276:immunology; D010949:Plasma / Q000276:immunology / Q000378:metabolism*; D020033:Protein Isoforms; D044967:Serum / Q000276:immunology / Q000378:metabolism*; D001613:beta 2-Microglobulin / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D006680:HLA Antigens; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I; D020033:Protein Isoforms; D001613:beta 2-Microglobulin