Use of the recombinant human thyrotropin receptor (TSH-R) expressed in mammalian cell lines to assay TSH-R autoantibodies.
We report on two assays for autoantibodies to the TSH-R which have been developed using materials from mammalian cells transfected with the cDNA for the human TSH-R. In the first, a particulate fraction has been prepared from COS cells, transiently expressing the human TSH-R and used in a radioreceptor assay in conjunction with bovine 125I-TSH. Immunoglobulins (IgGs) from patients with Graves' disease (n = 11) and idiopathic myxoedema (n = 2) have been used as competitors of 125I-TSH binding to the COS TSH-R membranes and the results have been compared with those obtained with a commercially available kit for measuring TSH-R autoantibodies, which uses solubilised porcine TSH-R. Both assays showed similar performance, being particularly sensitive to antibodies from patients with idiopathic myxoedema. In the second assay system we have used a CHO cloned cell line (JP26) stably transfected with the human TSH-R. A selection of IgG preparations from patients with Graves' disease and of six normal controls was used to test the ability of this cell line to detect thyroid stimulating immunoglobulins (TSAb) by increasing its cAMP production. The assay was performed under two conditions: in standard (isotonic) medium or in hypotonic medium. Freshly thawed human thyrocytes incubated in hypotonic medium served as a reference method. Only five patients scored positive when tested in the JP26 cell line under isotonic conditions. When the assay was performed in a hypotonic medium, a significant positive correlation was observed between the results given by JP26 cells and human thyrocytes.(ABSTRACT TRUNCATED AT 250 WORDS)
Ludgate
M
M
I.R.I.B.H.N., ULB, Hôpital Campus Erasme, Brussels, Belgium.
Perret
J
J
Parmentier
M
M
Gerard
C
C
Libert
F
F
Dumont
J E
JE
Vassart
G
G
D000818:Animals; D001323:Autoantibodies / Q000032:analysis* / Q000276:immunology; D002478:Cells, Cultured; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000096:biosynthesis; D004247:DNA / Q000235:genetics; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D006111:Graves Disease / Q000276:immunology*; D006801:Humans; D007038:Hypotonic Solutions / Q000494:pharmacology; D007074:Immunoglobulin G / Q000032:analysis; D018828:Immunoglobulins, Thyroid-Stimulating; D007668:Kidney; D009230:Myxedema / Q000276:immunology; D010053:Ovary; D011933:Reagent Kits, Diagnostic; D011989:Receptors, Thyrotropin / Q000235:genetics / Q000276:immunology; D011994:Recombinant Proteins; D013961:Thyroid Gland / Q000166:cytology
D001323:Autoantibodies; D007038:Hypotonic Solutions; D007074:Immunoglobulin G; D018828:Immunoglobulins, Thyroid-Stimulating; D011933:Reagent Kits, Diagnostic; D011989:Receptors, Thyrotropin; D011994:Recombinant Proteins; D004247:DNA; D000242:Cyclic AMP