A Sensitive Bioassay of Parathyroid Hormone in Plasma

Abstract

Understanding of calcium metabolism in health and disease has been retarded by the lack of an adequately sensitive bioassay of parathyroid hormone. The problem of dissociation of bioactivity and immunoactivity, well recognized for other polypeptide hormones, is exaggerated in the case of parathyroid hormone by the disproportionately long half-time in the circulation of the immunoreactive fragments. A new method of assaying the biological activity of parathyroid hormone in plasma has been developed, based on the cytochemical methods which have yielded highly sensitive bioassays of other polypeptide hormones. It depends on the stimulation of glucose 6-phosphate dehydrogenase activity in the distal convoluted tubules of segments of guinea-pig kidney maintained in vitro, and measured by microdensitometry. The limit of sensitivity of the assay is 5 fh/ml (bPTH); the index of precision is 0.09 +/- 0.04 (mean +/- SEM; n = 11).

Circulating Levels of Biologically Active and Immunoreactive Intact Parathyroid Hormone in Human Newborns

“…CBA was performed as reported previously by Chambers et al (20) and Fenton et al (26) with minor modifications (27). The assay procedure is based on the time-and dosedependent stimulation of G6PD activity by PTH in guinea pig renal cortical cells.…”

“…Segments were chilled by immersion in hexane on dry ice and stored at -70°C for 1-3 d. Unfixed frozen crystat sections (8 pm) were cut from just below the cortical surfaces at -12°C and then reacted for G6PD activity in 5 mM glucose 6-phosphate, 3 mM NADP, 0.67 mM phenazine methosulfate, 5 mM tetrazolium chloride, 10 mM potassium cyanide, and 20% polyvinyl alcohol in 0.05 M glycyl glycine buffer, pH 8.0. Juxtaglomerular DCT cells were identified by morphologic and, in parallel sections, cytochemical criteria (20). The intensity of the formazan reaction product within DCT cells was measured by microdensitometry (Zeiss) at 589 nm with an 8-pm mask.…”

“…Using the ultrasensitive CBA for PTH developed by Chambers (20), Care et al (21,22) showed that bioPTH in porcine fetal plasma exceeds paired maternal plasma bioPTH. Similarly, Allgrove et al (14) reported that human term umbilical cord plasma bioPTH is elevated 5-to 6-fold above paired maternal plasma bioPTH and significantly exceeds values determined in normocalcemic adults.…”

The role of parathyroid hormone‐related protein in calcium homeostasis in the fetal pig

“…Total PTH-like bioactivity was measured using a renal cytochemical assay (Chambers, Dunham, Zanelli, Parsons, Bitensky & Chayen, 1978) with human PTH (hPTH(1-84)) as standard. In this assay, bovine PTH(bPTH)(1-84) was equipotent with bPTH(I-34) on a molar basis (Loveridge, Caple, Rodda, Martin & Care, 1988).…”

Cytochemical bioassays: every dog has its day

“…The CBA for parathyroid hormone was developed by David Chambers 50 in response to a request from John Parsons at the National Institute for Medical Research. He had previously argued that the normal circulating concentration of PTH should be around 10-15 pg ml À1 but the current immunoassays at the time were recording values about 15-fold higher.…”