Development of a scintillation proximity assay for high-throughput measurement of intact parathyroid hormone.
A simple, high-throughput scintillation proximity assay (SPA) for parathyroid hormone (1-84) (PTH) has been developed. Fifteen commercially available N-terminal and six C-terminal anti-PTH antibodies were evaluated for detection of human PTH(1-84). Two C-terminal antibodies (CR1073M and 10-P55) gave the most consistent results. Using one of these antibodies (10-P55), an assay was developed with a sensitivity of 4 pg/ml for human and rat PTH(1-84). Porcine PTH(1-84) was not detectable. The intra-assay and inter-assay coefficients of variation for a 467 pg/ml sample were 6. 1 and 6.5%, respectively, and for a 21 pg/ml sample, 6.2 and 4.4%. Human PTH(1-34), while not detected in the assay, interfered with the detection of PTH(1-84). Smaller fragments [for example, human PTH(3-34)] and a C-terminal PTH fragment [PTH(53-84)] did not interfere in the assay. The procedure gave 106-110% recovery of human PTH(1-84) spiked into samples. Immunoreactive PTH concentrations in serum of rats administered EGTA were determined by SPA and by a commercially available PTH immunoassay. There was a good correlation between the two assays with significant increases in serum immunoreactive PTH concentrations at 15 and 30 min after EGTA injection and a rapid decrease to baseline values by 60 min. The SPA gives a high-throughput method for simply and accurately determining PTH(1-84) concentrations in serum.
Frolik
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CA
Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana, 46285, USA.
Black
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Chandrasekhar
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Adrian
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MD
D000818:Animals; D006801:Humans; D008297:Male; D010281:Parathyroid Hormone / Q000097:blood*; D051381:Rats; D017207:Rats, Sprague-Dawley; D015203:Reproducibility of Results; D012588:Scintillation Counting / Q000379:methods*; D012680:Sensitivity and Specificity
D010281:Parathyroid Hormone