Highly Specific and Sensitive Sandwich Enzyme Immunoassay for Human Thyroid-Stimulating Hormone (hTSH) in Serum Using Monoclonal Anti-hTSH β-Subunit IgG1-Coated Polystyrene Balls and Polyclonal Anti-hTSH β-Subunit Fab′-Horseradish Peroxidase Conjugate

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Abstract
A highly specific and sensitive sandwich enzyme immunoassay for human thyroid-stimulating hormone (hTSH) in serum is described. Monoclonal anti-hTSH β-subunit IgG1−coated polystyrene balls were incubated with serum samples and subsequently with affinity-purified polyclonal anti-hTSH β-subunit Fab′-horseradish peroxidase conjugate. There was no cross-reaction with 38 mIU/tube of human follicle-stimulating hormone, 40 mIU/tube of human luteinizing hormone or 10 IU/tube of human chorionic gonadotropin. The detection limit of hTSH was 0.3 nU/tube or 0.006 mU/1 of serum. Therefore, it was possible to measure very low levels of serum hTSH in pregnant women with Graves' disease. Serum hTSH levels in normal subjects and patients with Graves' disease were 2.45 ± 1.49 (SD) mU/1 (range : 0.61–6.09 mU/1) and 0.086 ± 0.060 (SD) mU/1 (range : < 0.006-0.18 mU/l), respectively. However, the present sandwich enzyme immunoassay was expensive, since affinity-purified polyclonal anti-hTSH β-subunit Fab′-peroxidase conjugate was prepared from rabbit anti-hTSH serum.