开发并验证一种 LC-MSMS 方法，用于定量检测干血斑中的肌酐含量

IF 3.1 4区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Journal of Mass Spectrometry and Advances in the Clinical Lab Pub Date : 2024-03-06 DOI:10.1016/j.jmsacl.2024.03.001

Carlos Torres , Rogers A. Muldrow , Anissa R. Naranjo , Steven W. Cotten , Christina C. Pierre , Dina N. Greene

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引用次数: 0

摘要

慢性肾脏病的筛查有赖于准确和精确的肌酐测量。传统上，使用高通量化学分析仪测量血清或血浆中的肌酐。不过，干血斑（DBS）也可用于改善检测通道。样本取自 6 毫米的干血点冲孔，在进行乙腈冲洗前在水中重组。所得上清液在注射前用 80:20 的乙腈：水稀释。使用等度梯度法鉴定肌酐，并使用 API 4000 三重四极杆质量分析仪进行检测。定量依赖于与基质匹配的校准物，其值与罗氏 Cobas 酶联免疫测定法一致。评估方法性能的验证研究包括精密度、线性度、准确度、方法比较、稳定性、干扰和基质效应。LC-MSMS 分析法在 0.3-20 mg/dL 范围内线性良好（y=1.02x-0.11；R=0.996）。使用横跨分析测量范围的基质匹配对照（n=4），精确度为 5.2-8.1%。LC-MSMS 结果与酶法检测（罗氏）结果一致，拟合直线方程为 y=0.956x-0.07 (R=0.995; n=173)。西门子和罗氏酶法测定与 DBS 肌酐浓度（n=40 成对静脉/DBS 采集）相关的准确度高于贝克曼-雅法测定（分别为 -2.5% 和 -0.8% 对 -6.3% 和 -4.1%）或 iSTAT（分别为 -28.4% 和 -27.1%）。准确性不受血细胞比容、血斑容量、IgG 或 IgA 过量或高甘油三酯血症的影响。没有观察到基质效应，提取和处理效率都很高。我们成功地开发了一种准确、精确的 LC-MSMS 方法，用于定量 DBS 中的肌酐。

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Development and validation of an LC-MSMS method to quantify creatinine from dried blood spots

Introduction

Screening for chronic kidney disease relies on accurate and precise creatinine measurements. Traditionally, creatinine is measured in serum or plasma using high-throughput chemistry analyzers. However, dried blood spots (DBS) can also be utilized to improve testing access.

Methods

Samples were obtained from a 6 mm DBS punch, which was reconstituted in water before undergoing an acetonitrile crash. The resulting supernatant was diluted using an 80:20 acetonitrile: water before injection. Creatinine was identified using an isocratic gradient, and detected using an API 4000 triple quadrupole mass analyzer. Quantification relied on matrix-matched calibrators, with values harmonized to the Roche Cobas enzymatic assay. Validation studies assessing method performance included precision, linearity, accuracy, method comparison, stability, interference, and matrix effects.

Results

The LC-MSMS assay was linear from 0.3 to 20 mg/dL (y = 1.02x-0.11; R² = 0.996). Precision ranged from 5.2 to 8.1 % using matrix-matched controls (n = 4) that spanned the analytical measurement range. LC-MSMS results corresponded to the enzymatic assay (Roche) with a fitted line equation of y = 0.956x–0.07 (R² = 0.995; n = 173). The Siemens and Roche enzymatic assays demonstrated higher accuracy in correlating to the DBS creatinine concentration (n = 40 paired venous/DBS collections) compared to the Beckman Jaffe assay (-2.5 % and −0.8 % versus −6.3 % and −4.1 %, respectively) or the iSTAT (-28.4 % and −27.1 %, respectively). Accuracy was unaffected by hematocrit, blood spot volume, excess IgG or IgA, or hypertriglyceridemia. No matrix effects were observed, and both extraction and processing efficiency were robust.

Ambient stability extended to at least 10 days, and exposure to extreme temperature did not affect the creatinine results.