Objective: The distinct biological properties of TSH-receptor (TSH-R) autoantibodies (TRAb) from patients with Graves' disease (GD) are yet unexplained on the molecular level. Here we compare serum concentration, affinity to the TSH-R, and binding sites on the TSH-R of stimulating (TSAb) and blocking (TBAb) TRAb.

Methods and patients: Four step affinity purification using human recombinant TSH-R was performed with 22 TRAb positive sera from GD patients (11 with only TSAb and 11 with only TBAb) and 5 control sera. Antibody concentration, TBII, TSAb/TBAb activity of the purified TRAb were assessed. Labelled purified TRAb were used for displacement studies with TRAb and additional 30 patients and 10 control sera.

Results: TRAb could be purified to 80-93% purity with recovery of the TBII, TSAb and TBAb activity. No TRAb could be purified from healthy individuals. The mean±SD concentration of TRAb was 17.3±5.4 µg/IU for the TSAb sera (range 9.6-25.9), and 18.2±8.5 µg/IU for the TBAb sera (range 4.6-29.2), respectively (p=0.79). Affinity was in the picomolar range for both TRAb subtypes with mean ± SD K_d for TSAb of 167±109 pM (60-410 pM), and 253±132 pM (80-410 pM) for TBAb, respectively (p=0.12). Purified and labelled TSAb and TBAb showed a very similar binding pattern to the TSH-R in displacement studies with unlabelled TSAb/TBAb or unpurified patients sera, indicating binding sites on the TSH-R in close proximity to each other.

Conclusion: TSAb and TBAb in the serum of patients with GD have similar characteristics. They are of low concentration with high affinity and have also similar binding epitopes on the TSH-R.