Thrombin-mediated degradation of parathyroid hormone in serum tubes

• Thrombin cleaves human PTH at the Arg 44–Asp 45 position
• Intact PTH tests are sandwich immunoassays, which may be affected by peptide cleavage
• Exogenous bovine thrombin in rapid serum tubes leads to time and temperature dependent decreases in measurable PTH
• The direct thrombin inhibitor hirudin reduces exogenous (and presumably endogenous) cleavage of PTH by thrombin

BackgroundIntact parathyroid hormone (PTH) tests are frequently sandwich immunoassays. Enzymes that cleave PTH may cause falsely lower PTH results. The objective of this study was to determine whether bovine thrombin in Becton Dickinson (BD) Vacutainer rapid serum tubes™ (RSTs) may lead to PTH results that are lower than in plasma separator tube™ (PST) or serum separator tube™ (SST) collections.MethodsTubes of blood (PST, SST, and RST) were collected from donors. PTH concentrations were measured on a Roche Cobas e602 analyzer in aliquots held at room temperature or 4 °C across time. Instrument comparison studies were also conducted on an Abbott Architect i1000SR and a Siemens Immulite 2000 XPi. Previously collected serum specimens were also incubated in exogenous bovine thrombin, the direct thrombin inhibitor hirudin, or both. Freshly collected RST specimens were also spiked with hirudin after clotting and centrifugation.ResultsSignificant decreases in PTH degradation rate constants were observed according to tube type, with degradation rates faster in RSTs than SSTs, and SSTs faster than PSTs. PTH degradation rate was temperature dependent. PTH decreases induced by exogenous bovine thrombin, as well as endogenous human thrombin, were reduced by hirudin.ConclusionsBovine thrombin is responsible for the decrease in PTH results observed in RSTs. Endogenous human thrombin, activated during clot formation, is likely responsible for the smaller decreases observed in non-RST sera versus plasma.