Reversed-phase high-performance liquid chromatography of human thyroid-stimulating hormone

Human pituitary thyroid-stimulating hormone (TSH) has been analysed by reversed-phase high-performance liquid chromatography on Aquapore RP-300, a 300-A pore-size short-alkyl-chain silica packing, in sodium chloride-containing buffers. At poH 2.0, two clearly resolved peaks were observed corresponding in retention time to alpha and beta subunit of TSH. With increasing pH, the resolution between these peaks was lost. At pH 7.0 a single major peak was observed, co-eluting with immunoreactive TSH, which could be resolved from beta-subunit. In vivo TSH bioactivity was recovered at pH 7.0 in 58% yield. High-performance liquid chromatography of a sample of hypothyroid serum yielded a major peak of TSH immunoreactivity corresponding in retention time to that observed for purified TSH.