title: A direct binding assay for thyrotropin receptor autoantibodies.
abstract: There is, at present, no assay in clinical use for the direct assay of autoantibody binding to the thyrotropin receptor (TSHR). We now describe a direct thyrotropin receptor autoantibody binding assay (DTAb) using a secreted form of the TSHR ectodomain (TSHR-289) without the need for antigen purification. The assay compensates for the low TSHR autoantibody concentration in serum by capturing a relatively large amount of patient immunoglobulin G (IgG) on high-capacity beads, a reversal of standard methods that typically first immobilize antigen. TSHR-289 captured by Graves' IgG was detected in a colorimetric reaction using a biotinylated murine monoclonal antibody to the poly-histidine tail engineered into the antigen. By this approach, sera from 11 normal individuals provided a mean optical density (OD) value of 0.20 +/- 0.08 SD (range 0.06-0.33). Of 38 sera from unselected patients with a history of Graves' disease (untreated and treated), 29 (76%) generated OD values > 0.37 (2 SD above the mean for the normal sera), the highest being OD 1.38. Surprisingly, 3 of 13 (23%) sera from TPO autoantibody-positive patients with Hashimoto's thyroiditis also provided values > 2 SD above the normal sera. The extent of direct autoantibody binding to the TSHR correlated closely with the thyrotropin binding inhibition (TBI) values (r = 0.881; p < 0.001). One serum was clearly positive in only the direct binding assay and another in only the TBI assay. The data obtained with the direct binding assay correlated less well with the thyroid-stimulating antibody (TSAb) assay (r = 0.582; p < 0.001). In summary, we describe a new direct DTAb assay that correlates more closely with the TBI than with the TSI assays. Future studies in a large series of clinically defined patients will be needed to evaluate the clinical utility of the DTAb assay.
authors.affiliation: Autoimmune Disease Unit, Cedars-Sinai Research Institute and School of Medicine, University of California, Los Angeles 90048, USA.
authors.forename: G D
authors.initials: GD
authors.lastname: Chazenbalk
authors.forename: P
authors.initials: P
authors.lastname: Pichurin
authors.forename: S M
authors.initials: SM
authors.lastname: McLachlan
authors.forename: B
authors.initials: B
authors.lastname: Rapoport
mesh_terms: D001323:Autoantibodies / Q000097:blood*; D006801:Humans; D011989:Receptors, Thyrotropin / Q000276:immunology*
chemical_list: D001323:Autoantibodies; D011989:Receptors, Thyrotropin