Department of Endocrine Substance Analysis Center (ESAC), Yongin, Korea.
GC Labs, Yongin, Korea.
Ann Lab Med. 2025 Jan 1;45(1):62-69. doi: 10.3343/alm.2024.0072. Epub 2024 Aug 23.
Mass spectrometry (MS) methods exhibit higher accuracy and comparability in measuring serum C-peptide concentrations than immunoassays. We developed and validated a novel isotope dilution-ultraperformance liquid chromatography-tandem MS (ID-UPLC-MS/MS) assay to measure serum C-peptide concentrations.
Sample pretreatment involved solid-phase extraction, ion-exchange solid-phase extraction, and derivatization with 6-aminoquinolyl-N-hydroxysuccinimidylcarbamate (Cayman Chemical, Ann Arbor, Michigan, USA). We used an ExionLC UPLC system (Sciex, Framingham, MA, USA) and a Sciex Triple Quad 6500+ MS/MS system (Sciex) for electrospray ionization in positive-ion mode with multiple charge states of [M+3H]3+ and multiple reaction monitoring transitions. The total run time was 50 mins, and the flow rate was 0.20 mL/min. We evaluated the precision, trueness, linearity, lower limit of quantitation (LLOQ), carryover, and matrix effects. Method comparison with electrochemiluminescence immunoassay (ECLIA) was performed in 138 clinical specimens.
The intra- and inter-run precision coefficients of variation were <5% and the bias values for trueness were <4%, which were all acceptable. The verified linear interval was 0.050-15 ng/mL, and the LLOQ was 0.050 ng/mL. No significant carryover or matrix effects were observed. The correlation between this ID-UPLC-MS/MS method and ECLIA was good (R=0.995, slope=1.564); however, the ECLIA showed a positive bias (51.8%).
The developed ID-UPLC-MS/MS assay shows acceptable performance in measuring serum C-peptide concentrations. This will be useful in situations requiring accurate measurement of serum C-peptide in clinical laboratories.
质谱（MS）方法在测量血清 C 肽浓度方面比免疫测定具有更高的准确性和可比性。我们开发并验证了一种新的同位素稀释-超高效液相色谱-串联质谱（ID-UPLC-MS/MS）测定法来测量血清 C 肽浓度。
样品预处理包括固相萃取、离子交换固相萃取和 6-氨基喹啉基-N-羟基琥珀酰亚胺基碳酸酯（Cayman Chemical，密歇根州安阿伯，美国）衍生化。我们使用 ExionLC UPLC 系统（Sciex，马萨诸塞州弗雷明汉，美国）和 Sciex Triple Quad 6500+ MS/MS 系统（Sciex）进行电喷雾正离子模式下的多电荷状态 [M+3H]3+和多重反应监测转换。总运行时间为 50 分钟，流速为 0.20 mL/min。我们评估了精密度、准确度、线性、定量下限（LLOQ）、交叉污染和基质效应。在 138 份临床标本中进行了与电化学发光免疫测定（ECLIA）的方法比较。
日内和日间精密度变异系数<5%，准确度偏差值<4%，均在可接受范围内。验证的线性区间为 0.050-15 ng/mL，LLOQ 为 0.050 ng/mL。未观察到明显的交叉污染或基质效应。该 ID-UPLC-MS/MS 方法与 ECLIA 之间的相关性良好（R=0.995，斜率=1.564）；然而，ECLIA 显示出正偏差（51.8%）。
所开发的 ID-UPLC-MS/MS 测定法在测量血清 C 肽浓度方面表现出可接受的性能。这将在临床实验室需要准确测量血清 C 肽的情况下非常有用。
