Application of a Bioassay with CHO Cells for the Routine Detection of Stimulating and Blocking Autoantibodies to the TSH-Receptor

Abstract

The importance of bioassays measuring stimulating and blocking autoantibodies to the TSH-receptor (TSH-R) by their effect on cAMP production in CHO cells transfected with the recombinant TSH-R is increasingly recognized. The standard technique for this bioassay is cumbersome, as it involves purification of serum IgG with polyethylene glycol (PEG) and resuspension in hypotonic buffer. We have therefore established a simpler approach for the detection of stimulating and blocking autoantibodies using JP09 CHO cells and unfractionated human serum. The cAMP concentration was measured by a highly sensitive commercial radioimmuno assay. Thyroid stimulating autoantibodies (TSAb) were present in 107 out of 126 patients with Graves' disease (85%) and in 4 out of 40 patients with Hashimoto's thyroiditis (10%). Specificity was confirmed by the fact that only 1 patient with insulin dependent diabetes mellitus (IDDM) out of 64 patients with different non-thyroid autoimmune disorders (46 with IDDM, 10 with stiff man syndrome and 8 with rheumatoid arthritis) and 2 out of 100 healthy controls (2%) were positive in this assay. In the subgroup of hyperthyroid Graves' disease patients 76 out of 83 (92%) had TSAb and the same number had TSH binding inhibiting immunoglobulin (TBII), as assessed by the commercial TRAK assay. Although both antibody types showed only a weak correlation (r = 0.30), a combination of TSAb and TBII detected 98% of all Graves' patients and 99% of the hyperthyroid subgroup. Thyroid blocking autoantibodies (TBAb) were measured in 4 out of 24 TSAb negative patients with Graves' disease (17%), who were hypothyroid and positive for TBII. A comparison of our bioassay with the standard bioassay using PEG precipitation showed a good correlation (r = 0.76,p < 0.001), demonstrating the feasibility of the simplified assay for the routine detection of TSAb and TBAb in Graves' disease.