title: Typing of serum-soluble HLA-B27 antigen by ELISA.
abstract: An ELISA using serum as soluble HLA antigen source was developed for HLA-B27 typing. Two sandwich assays were run in parallel. The first assay utilized a monoclonal antibody (mAb) reacting with a determinant expressed by both HLA-B7 and B27 antigens; the other assay utilized a mAb reactive with HLA-B7 antigens but not with HLA-B27 antigens. After incubation with serum samples, bound HLA antigen was detected using an anti-beta 2m antibody conjugated to peroxidase and a chromogenic substrate. Absorbance of each well was measured at 490 nm. Based on analysis of absorbances obtained with panels of specimens of known HLA phenotypes, a mathematical algorithm was developed to derive the specimen HLA-B27 phenotype from its ELISA absorbance values. Despite the lack of monospecific mAb, an accurate HLA-B27 typing was possible. 362 specimens (including 151 HLA-B27-positive) were tested. Agreement between microlymphocytotoxicity and ELISA was 99.2%. No correlation between the level of HLA-B27 antigen reactivity and the amount of total HLA class I antigen in serum was observed. This report demonstrates the possibility of using serum-soluble HLA antigen and ELISA technology for histocompatibility testing. The assay offers several significant advantages over microlymphocytotoxicity: no need for cell preparation, batch testing capabilities and objective, reproducible interpretation of results.
authors.affiliation: SangStat Medical Corporation, Menlo Park, California.
authors.forename: P
authors.initials: P
authors.lastname: Pouletty
authors.forename: C
authors.initials: C
authors.lastname: Chang
authors.forename: J
authors.initials: J
authors.lastname: Kalil
authors.forename: E
authors.initials: E
authors.lastname: Atwood
authors.forename: S
authors.initials: S
authors.lastname: Ferrone
authors.forename: B
authors.initials: B
authors.lastname: Shimizu
authors.forename: W
authors.initials: W
authors.lastname: Howson
authors.forename: R
authors.initials: R
authors.lastname: Mazaheri
authors.forename: B
authors.initials: B
authors.lastname: Del Villano
authors.forename: C
authors.initials: C
authors.lastname: Grumet
mesh_terms: D000465:Algorithms; D000911:Antibodies, Monoclonal / Q000032:analysis / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D015796:HLA-B27 Antigen / Q000032:analysis / Q000145:classification* / Q000276:immunology; D015793:HLA-B7 Antigen / Q000032:analysis / Q000276:immunology; D006801:Humans; D010641:Phenotype; D015203:Reproducibility of Results; D012984:Software
chemical_list: D000911:Antibodies, Monoclonal; D015796:HLA-B27 Antigen; D015793:HLA-B7 Antigen