Background Parathyroid hormone (PTH) measurement is important for patients with disorders of calcium metabolism, including those needing bone-turnover monitoring due to chronic kidney disease–mineral bone disorder. There are currently 2 generations of PTH immunoassays on the market, both having cross-reactivity issues and lacking standardization. Therefore, we developed an LC-MS/MS higher-order method for PTH analysis. Methods The method was calibrated against the international standard for 1-84 PTH (WHO 95/646). Antibody-free sample preparation with the addition of an isotope-labeled internal standard was performed by solid-phase extraction. Extracts were analyzed by LC-MS/MS. EDTA-K2 plasma was used throughout the development and validation. Bias and uncertainty sources were tested according to ISO 15193. Clinical Laboratory Standards Institute guidelines and reference measurement procedures were consulted for the design of the validation. Patient samples and external quality controls were compared between LC-MS/MS and 2 third-generation immunoassays. Results The method was validated for 1-84 PTH from 5.7 to 872.6 pg/mL. The interassay imprecision was between 1.2% and 3.9%, and the accuracy ranged from 96.2% to 103.2%. The measurement uncertainty was <5.6%. The comparison between LC-MS/MS and the immunoassays showed a proportional bias but moderate to substantial correlation between methods. Conclusions This LC-MS/MS method, which is independent of antibodies, is suitable for a wide range of PTH concentrations. The obtained analytical performance specifications demonstrate that development of a reference measurement procedure will be possible once a higher order reference standard is available.

中文翻译：

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使用固相萃取定量 1-84 甲状旁腺激素的 LC-MS/MS 方法的验证：寻找候选参考测量程序


背景 甲状旁腺激素 (PTH) 测量对于钙代谢紊乱的患者非常重要，包括因慢性肾病 - 矿物质性骨病而需要骨转换监测的患者。目前市场上有2代PTH免疫检测，均存在交叉反应问题且缺乏标准化。因此，我们开发了一种用于 PTH 分析的 LC-MS/MS 高阶方法。方法 该方法根据 1-84 PTH 国际标准 (WHO 95/646) 进行校准。通过固相萃取进行无抗体样品制备并添加同位素标记的内标。通过 LC-MS/MS 分析提取物。 EDTA-K2 血浆用于整个开发和验证。根据 ISO 15193 测试偏差和不确定性来源。验证设计参考了临床实验室标准协会指南和参考测量程序。对 LC-MS/MS 和 2 个第三代免疫测定法之间的患者样本和外部质量控制进行了比较。结果 该方法针对 5.7 至 872.6 pg/mL 的 1-84 PTH 进行了验证。批间不精密度为1.2%至3.9%，准确度为96.2%至103.2%。测量不确定度为<5.6%。 LC-MS/MS 和免疫测定之间的比较显示出比例偏差，但方法之间存在中度至显着相关性。结论 这种 LC-MS/MS 方法不依赖于抗体，适用于较宽范围的 PTH 浓度。获得的分析性能规格表明，一旦获得更高阶的参考标准，就可以开发参考测量程序。