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A Coated Tube Assay for the Detection of Blocking Thyrotropin Receptor Autoantibodies

MiLo GmbH (W.B.M.), Bioassays GmbH (C.L.), and Research Department (A.B., N.G.M.), B.R.A.H.M.S. AG, Biotechnology Center Hennigsdorf/Berlin, D-16761 Hennigsdorf, Germany

Address all correspondence and requests for reprints to: Dr. Nils G. Morgenthaler, Research Department, B.R.A.H.M.S. AG, Biotechnology Center Hennigsdorf/Berlin, Neuendorfstr. 25, D-16761 Hennigsdorf, Germany. E-mail: n.morgenthaler{at}brahms.de.

We developed a coated tube assay to discriminate TSH-receptor-stimulating autoantibodies [thyroid-stimulating antibodies (TSAb)] from those autoantibodies blocking TSH binding without intrinsic activation [thyroid-blocking antibodies (TBAb)]. The wild-type TSH receptor in the TSH binding-inhibitory assay was exchanged for a chimeric receptor where a TSAb epitope (amino acids 8–165) was replaced by comparable LH-R residues. Binding of ¹²⁵I-labeled TSH to this chimera could be inhibited by sera containing TBAb up to 95%. Sera from 316 patients with Graves’ disease and 17 with autoimmune thyroid disease were grouped according to their bioassay activity. At the decision threshold, the chimera A assay had a sensitivity of 78.0% for TBAb with a specificity of 90.2%. In detail, 19 of 22 (86.4%) TBAb sera and 15 of 23 (65.2%) TSAb/TBAb sera were positive but only 32 of 216 (14.0%) TSAb sera and 5 of 72 (6.9%) bioassay negative sera. There was a weak but significant positive correlation (r = 0.46) between the chimera assay and the bioassay for TBAb. This is the first report of a coated tube assay for the determination of TBAb employing an adaptation of the TSH binding-inhibitory format, which could be a useful alternative to the bioassay.

This work was supported by a grant from the European Union and the State of Brandenburg (Verfahrensinnovation no. 80084492) to MiLo GmbH.

Abbreviations: CI, Confidence interval; GD, Graves’ disease; LH-CG, lutropin/choriogonadotropin; ROC, receiver-operating characteristic; TBAb, thyroid-blocking antibodies; RLU, relative light units; TBII, TSH binding inhibition; TRAb, TSH-R autoantibodies; TSAb, thyroid-stimulating antibodies; TSH-R, TSH receptor; WT, wild-type.

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