Journal of Korean Endocrine Society 1998;13(2):167-180.
Published online January 1, 2001.
Usefulness of Immunoglobulin Fraction Precipitated with Polyethylene Glycol in Assay for TSH Receptor Antibodies using Chinese Hamster Overy Cells Expressing Human TSH Receptors.
Won Bae Kim, Hyun Kyung Chung, Chang Soon Koh, Chang Hoon Yim, Do Joon Park, Bo Yeon Cho, Hong Gyu Lee
Graves' disease and primary myxedema are thought to be caused by the action of TSH receptor autoantibodies(thyroid stimulating antibody; TSAb & thyroid stimulation blocking antibody; TSBAb). Thus, detection of these antibodies is crucial in diagnosis and in follow up of those patients. Recently, a sensitive method using human TSH receptor transfected Chinese Hamster Ovary(CHO) cells has been developed. However, the complexity of IgG purification procedure is considered as a limitation for its clinical application as a routine test. The aim of this study is to determine whether polyethylene glycol(PEG)-precipitated immunogiobuIin fraction could substitute for purified IgG. METHODS: We developed optimal conditions for TSAb and TSBAb assays using crude, PEG precipitated immunoglobulin fraction; and evaluated the correlation of TSAb and TSBAb activities between thase measured using crude immunoglobulin fraction and purified IgG to clarify the usefulness of PEG-precipitated immunoglobulin fraction. TSH receptor expressing wild type CHO cells were used in TSAb and CHO cells expressing chimeric TSH receptor(Mc2; 90-165 amino acid residues were substituted by those of rat LH/CG receptar) were used in TSBAb assay to minimize the possible disturbing effects of TSAb in serum. RESULTS: The optimal serum amount for TSAb and TSBAb assay using PEG-precipitated immunoglobulin fraction were 250mL serum equivalent/well and 50mL serum equivalent/well, respectively. The optimal incubation time for both assays were 2 homs, and aptimal ccrncentration of bTSH for TSBAb assay was 0.1U/L. TSAb activities measured with PEG-precipitated immunoglobulin were significantly correlated with those measured with purified IgG in 26 patients with Graves diseases(r=0.93, p<0.001). Although TSBAb activities measured using PEG-precipitated imrnunoglobulin were conelated with those measured using purified IgG in 20 patients with primary myxedema(r=0.86, p<0.001), the positive rate in TSBAb assay using PEG-precipitated immunoglobulin was lower than that of usmg purified IgG(20% v.s. 65%) because of negative conversion of TSBAb activities in samples with weakly positive TSBAb activities measured using purified IgG. CONCLUSION: PEG-precipitated immunoglobulin fraction could be used instead of purified IgG in TSAb assay using hTSHR-tranasfected wild type CHO cells with equal sensitivity and specificity. This simple and practical TSAb assay using PEG-precipitated immunoglobulin in hTSHR-transfected CHO cells would be useful in clinica1 practiee.
Key Words: TSAb, TSBAb, hTSHR-transfected CHO cell, PEG-precipitated immunoglobuhn

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