Establishment of Time-Resolved Fluorescence Immunoassay for Thyrotropin Receptor Antibodies and Clinical Application.

Thyroid-stimulating hormone receptor antibody (TRAb) is a specific marker for Graves' disease (GD) and the measurement of which can improve the accuracy of GD diagnosis. Current detection methods utilize porcine-derived polyclonal-TRAb, which is unstable and is a source of significant inter-assay variability. This study aims to establish a time-resolved fluorescence immunoassay (TRFIA) method based on stable source of recombinant human TSHR and TRAb for the detection of serum TRAb. The neutralization inhibition method was used in this study. The specific binding of goat-anti-mouse IgG and chimeric construct Fc fragment human TSHR were immobilized on the microplate, and serum TRAb was detected by the competition between Eu-labeled human TRAb and the serum TRAb. The TRAb-TRFIA has a wide linear range (0.081-50 IU/L). The intra-assay precision was 2.17-5.41% (< 10%), and the inter-assay precision was 5.75-9.58% (< 15%). No cross-reactivity was observed between TRAb and recombinant human IgG1, IgG2, IgG3, or IgG4. The recovery rate was 106.37%. The TRAb-TRFIA method was significantly correlated with the Roche chemiluminescence method (R = 0.9130). TRFIA based on recombinant human TSHR and TRAb with high stability, strong specificity, high sensitivity and wide linear range was established for the detection of TRAb, which can be used for clinical detection.