Sohn Areum, Kim Hyunsoo, Yu Su Jong, Yoon Jung-Hwan, Kim Youngsoo
Department of Biomedical Sciences, Seoul National University College of Medicine, Yongon-Dong, Seoul, 110-799, Korea.
Department of Biomedical Engineering, Seoul National University College of Medicine, Yongon-Dong, Seoul, 110-799, Korea.
Anal Bioanal Chem. 2017 Apr;409(11):2829-2838. doi: 10.1007/s00216-017-0227-8. Epub 2017 Feb 6.
Prothrombin induced by vitamin K absence-II (PIVKA-II) is an effective tumor marker for hepatocellular carcinoma (HCC). We have developed a novel targeted mass spectrometric (MS) assay for quantifying PIVKA-II in human serum. The ideal signature peptide was selected to measure PIVKA-II concentrations on a triple quadrupole (QqQ) mass spectrometer, and the chromatography gradient was optimized for the peptide separation to minimize elution interference. Using multiple reaction monitoring-mass spectrometry (MRM-MS), good linearity (R = 0.9988) was obtained for PIVKA-II over a range of 3 orders. We achieved a limit of detection (LOD) of 0.45 nM (31.72 ng/mL), a limit of quantification (LOQ) of 0.93 nM (65.31 ng/mL), a lower limit of quantification (LLOQ) of 0.49 nM (34.32 ng/mL), and an upper limit of quantification (ULOQ) of 1000.00 nM (70,037.00 ng/mL). The intra-day and inter-day precisions were within ±14.96%, and the accuracy ranged from 87.66 to 114.29% for QC samples at four concentrations. Compared with an established immunoassay, the correlation (R = 0.8335) was good for the measurements of PIVKA-II concentrations. This method was successfully applied to the analysis of clinical samples for normal control (n = 50), chronic hepatitis (n = 50), liver cirrhosis (n = 50), HCC (n = 50), and recovery (n = 50) serum. Graphical Abstract MRM-MS assay development for determining concentration of PIVKA-II in serum and a comparison between MRM-MS assay and immunoassay with high correlation.
维生素K缺乏诱导蛋白II（PIVKA-II）是肝细胞癌（HCC）的一种有效肿瘤标志物。我们开发了一种新型靶向质谱（MS）分析法，用于定量人血清中的PIVKA-II。选择理想的特征肽在三重四极杆（QqQ）质谱仪上测量PIVKA-II浓度，并优化色谱梯度以进行肽分离，从而将洗脱干扰降至最低。使用多反应监测质谱法（MRM-MS），PIVKA-II在3个数量级范围内具有良好的线性（R = 0.9988）。我们实现了0.45 nM（31.72 ng/mL）的检测限（LOD）、0.93 nM（65.31 ng/mL）的定量限（LOQ）、0.49 nM（34.32 ng/mL）的最低定量限（LLOQ）和1000.00 nM（70,037.00 ng/mL）的最高定量限（ULOQ）。日内和日间精密度在±14.96%以内，四种浓度的质量控制（QC）样品的准确度范围为87.66%至114.29%。与既定的免疫分析法相比，PIVKA-II浓度测量的相关性良好（R = 0.8335）。该方法成功应用于正常对照（n = 50）、慢性肝炎（n = 50）、肝硬化（n = 50）、HCC（n = 50）和康复期（n = 50）血清的临床样本分析。图形摘要用于测定血清中PIVKA-II浓度的MRM-MS分析法开发以及MRM-MS分析法与具有高相关性的免疫分析法之间的比较。
