Immunoreactivity of plasma parathyrin-related peptide: three region-specific radioimmunoassays and a two-site immunoradiometric assay compared

Abstract

We measured parathyrin (parathyroid hormone)-related peptide (PTHRP) in plasma by three region-specific RIAs and compared them with an established two-site immunoradiometric assay (IRMA) of PTHRP1-86 in samples from control subjects and from patients with primary hyperparathyroidism (PH) and humoral hypercalcemia of malignancy (HHM). The two direct RIAs of PTHRP1-34 and PTHRP37-67 were specific for regions 9-18 and 52-61, respectively. In the extraction RIA of PTHRP1-34 we used an affinity gel containing a monoclonal antibody specific for the 17-27 sequence; cross-reacting PTHRP species eluted from the gel were assayed by the RIA of PTHRP1-34. PTHRP1-86 plasma concentrations by IRMA were less than 0.23 pmol/L in control subjects and patients with PH, and were significantly increased in patients with HHM (mean 6.1 pmol/L, P less than 0.001). In contrast, plasma PTHRP1-34 concentrations were not significantly different in the three groups; in HHM patients, the mean was 190 pmol/L. Plasma PTHRP37-67 concentrations were similar in control and PH groups and, although significantly increased in HHM patients (mean 440 pmol/L, P less than 0.002), showed poor diagnostic discrimination. PTHRP1-34 concentrations after affinity extraction of plasma were also significantly higher in HHM patients (mean 10.7 pmol/L, P less than 0.001), but showed incomplete diagnostic discrimination. We conclude that the diagnostic utility of the direct RIAs for quantifying PTHRP is markedly inferior to the IRMA of PTHRP1-86.

Circulating levels of midregional parathyroid hormone‐related protein in hypercalcaemia of malignancy

“…This is comparable with recent results of other assays against different sequences ofPTHrP. With few exceptions (Grill et al, 1990), measuring amino terminal PTHrP in plasma was not possible by direct RIA and required laborious preceding extraction steps, such as immunoaffinity chromatography (Budayr et al, 1989;Ratcliffe et al, 1991b) or C-2 or C-18 extraction cartridges (Kao et af., 1990). With these methods, Budayr et al (1989) found elevated levels in 33 of 48 (69%) patients with a solid tumour and hypercalcaemia, and Kao et al (1990) in 12 of 25 patients (48%).…”

“…Very recently, Ratcliffe et al, have developed an RIA against PTHrP(37-67). The antiserum ofqhis assay was thought to recognize the region spanning residues 52-61 and was used successfully in a twosite assay (Ratcliffe et al, 1991b). The RIA against PTHrP(37-67), however, detected much higher concentrations of PTHrP in normals (mean: 320 pmol/l) than our RIA, although the antiserum was directed against a nearby epitope.…”

“…Supernatants from cell cultures of the hypercalcaemiainducing Walker carcinosarcoma 256 (Scharla et al, 1991a) contained 150-800 pmol/l of midregional PTHrP. In four samples of human breast milk from lactating women, the concentrations were between 12000 and 90000 pmol/l.…”

“…Since the known PTH-like biological activity of PTHrP is contained within the first 34 amino acids, most radioimmunoassays (RIA) have been directed against residues 1 to 34-40 (Budayr et al, 1989;Grill et al, 1990;Henderson et al, 1990;Kao et al, 1990;Ratcliffe & Ratcliffe, 1990; or against larger fragments containing amino terminal PTHrP (Burtis et al, 1990;. Although amino terminal PTHrP has been measured in plasma by direct RIA (Henderson et al, 1990;Ratcliffe et a[., 1991b;Scharla et al, 1991b), prior extraction steps were usually required to obtain clinically useful results (Budayr et al, 1989;Kao et al, 1990). This situation resembles the difficulties with assays for amino terminal PTH and may be complicated by the very low levels (< 2.5 pmol/l) of amino terminal PTHrP extracted by immunoaffinity chromatography from normal serum (Budayr et al, 1989).…”

The Relaxant Effects of Parathyroid Hormone(1‐34) and Parathyroid Hormone‐Related Protein(1‐34) on Ovine Reticulo‐Ruminal Smooth Muscle in Vivo

“…The plasma levels of PTHrP(1-34) were measured with an in-house radioimmunoassay using an antiserum raised in rabbits to hPTHrP(1-34) which is specific for the 9-18 region. The validation of this assay is described by Ratcliffe et al (1991).…”