Hyperthyroidism in Graves disease is due to TSH receptor (TSHR) autoantibodies (TRAb) directed against the TSH binding pocket. Detection of TRAb in clinical rou-tine is performed by 2nd gen assays using human or porcine TSHR. Such TSH bin- ding inhibition immunoglobulin (TBII) assays determine the ability of TRAb to inhi- bit the binding of labelled TSH to immobilized TSHR. TRAb are present in low con- centrations and mostly recognize conformational epitopes. The conformational structure of the TSHR is sensitive to thermal stress. Incubation times of current as- says must be for a minimum of 2 h to obtain good sensitivity. Due to these tech- nical difficulties no rapid automated TRAb assay could be developed so far.
We report on the 1st automated TRAb assay on the Elecsys/ cobas e immuno- assay platform. TRAb inhibit the binding of a human monoclonal thyroid stimula- ting autoantibody (M22; labelled with a ruthenium complex) to preformed immuno- complexes based on aggregates of solubilised multiple porcine TSHR (pTSHR) and a mouse monoclonal capture antibody (4E31 IgG; labelled with biotin). 4E31 does not interfere with the binding of TRAb and M22 to the pTSHR. The lyophili- zed immunocomplexes are stabilized after reconstitution by chemical chaperones. The assay uses a delayed competitive test principle. Total assay time is 3×9 min only. Firstly, sample TRAb if present bind to the multiple TSHR binding sites. Secondly, TRAb are allowed to interact with the TSHR further. Thirdly, labelled M22 and streptavidin-coated microparticles are added. M22 binds to still free bin- ding sites. Immunocomplexes are bound to the solid phase via interaction of biotin and streptavidin. Assay calibration is against the NIBSC 90/672. Its measuring range extends from 0.340 IU/l. The functional sensitivity is ~0.9 IU/l.
The automated Elecsys Anti-TSHR correlates well with current TBII assays and provides clinical performance and total precision which will meet customer needs.
03 - 07 May 2008
European Society of Endocrinology