Starting with a previously isolated cDNA for human thyrotropin receptor (TSHR), we established a transformed myeloma cell line, SP56, which expresses human TSHR on its cell surface. Binding analysis showed that SP56 bears 1.1×106TSHR per cell with a Kd of 2.2×10-10M. Using the purified cellular membrane, we established a TSH binding inhibition immunoglobulin (TBII) assay for autoantibodies against TSHR. We compared it with the TBII assay utilizing porcine thyroid membranes expressing porcine TSHR, which has been widely used for TBII assay, by using 96 serum samples from patients with autoimmune thyroid disease and normal individuals. Our TBII assay was more sensitive than the one using porcine TSHR: of 38 sera of patients which were judged negative for autoantibodies to TSHR(TBII value below 10%) by the latter assay, 28 were positive (above 20%) in our assay. By using a perfusion culture system, we obtained as many as 3×1010SP56 cells, from which 3, 450 mg protein of the membrane could be purified; this is sufficient for 15,000 assays. The results indicate that the membrane of the myeloma cell line SP56 is more suitable for use in the TBII assay than the porcine thyroid membrane, in terms of sensitivity to autoantibodies against TSHR in human sera.
|ジャーナル||Journal of biochemistry|
|出版ステータス||Published - 1995 8月|
ASJC Scopus subject areas