An accurate measurement method for serum homocysteine measurement by ID-LC/MS/MS and the application of external quality assessment

IF 3.2 3区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Clinica Chimica Acta Pub Date : 2024-11-01 DOI:10.1016/j.cca.2024.120021

Mo Wang , Jie Shi , Yichuan Song, Shunli Zhang, Rui Zhang

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引用次数: 0

Abstract

Background

Serum homocysteine (Hcy) measurement accuracy is essential for detecting and diagnosing cardio-cerebrovascular illnesses. Although many different methods have been developed to determine the concentration of Hcy, those different assays showed nonequivalent results. This study aimed to develop an accurate and precise isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC/MS/MS) method for serum homocysteine (Hcy) and assign values for reference materials used in external quality assessment (EQA) programs.

Methods

Different concentrations of homocysteine calibration solutions were prepared with d4-Hcy as the internal standard. This method employed DTT to reduce protein-bound Hcy, followed by protein precipitation, and gradient elution on a Supelcosil LC-CN column for chromatographic separation, and multiple reaction monitor (MRM) mode with electrospray ionization (ESI) for mass spectrometric detection. After optimized ID-LC/MS/MS parameters, imprecision, trueness, the limit of quantification (LoQ), the limit of detection (LoD), and measurement uncertainty were evaluated to check the methodological performance. The established method was employed in assigning values to EQA samples, which were sent to 63 clinical laboratories in Beijing to measure Hcy.

Results

At 10.69,15.99, and 37.80 μmol/L levels, the present method demonstrated analytical imprecision of 0.57 %, 0.65 %, and 0.57 %, and recoveries of 99.67 % to 100.21 %, respectively. The bias between the target values of GBW (Guojia Biaozhun Wuzhi) were − 0.79 % to 0.62 %. The LoQ and LoD for Hcy were 0.36 nmol/L and 0.27 nmol/L. The method had an uncertainty (U 95 %) of 1.34 % to 1.48 %. For the three levels of EQA samples, the percentage of laboratories meeting the trueness evaluation criteria (±10 %) was 81.67 %, 83.33 %, and 71.67 % respectively.

Conclusions

With optimal method precision and trueness, the ID-LC/MS/MS method to measure serum Hcy can be used for value assignment of EQA samples, which can provide reliable data for monitoring the accuracy of clinical laboratory for Hcy measurement.

查看原文本刊更多论文

利用 ID-LC/MS/MS 测量血清同型半胱氨酸的精确测量方法及外部质量评估的应用。

背景：血清同型半胱氨酸（Hcy）测量的准确性对于检测和诊断心脑血管疾病至关重要。尽管目前已开发出许多不同的方法来测定 Hcy 的浓度，但这些不同的测定方法显示出的结果并不一致。本研究旨在开发一种准确、精确的同位素稀释液相色谱-串联质谱法（ID-LC/MS/MS）测定血清同型半胱氨酸（Hcy）的方法，并为外部质量评估（EQA）计划中使用的参考材料赋值：以 d4-Hcy 为内标，制备不同浓度的同型半胱氨酸校准溶液。该方法采用 DTT 还原蛋白质结合的 Hcy，然后沉淀蛋白质，在 Supelcosil LC-CN 色谱柱上梯度洗脱进行色谱分离，并采用电喷雾离子化 (ESI) 多反应监测器 (MRM) 模式进行质谱检测。在对 ID-LC/MS/MS 参数进行优化后，对不精确度、真实度、定量限（LoQ）、检出限（LoD）和测量不确定性进行了评估，以检查方法的性能。采用既定方法对送往北京 63 家临床实验室测定 Hcy 的 EQA 样品进行赋值：结果：在 10.69、15.99 和 37.80 μmol/L 水平下，该方法的分析不精密度分别为 0.57 %、0.65 % 和 0.57 %，回收率为 99.67 % 至 100.21 %。与GBW（国药准字）目标值之间的偏差为- 0.79 %至0.62 %。Hcy的LoQ和LoD分别为0.36 nmol/L和0.27 nmol/L。该方法的不确定度（U 95 %）为 1.34 % 至 1.48 %。在三个等级的 EQA 样品中，符合真实度评价标准（±10 %）的实验室比例分别为 81.67 %、83.33 % 和 71.67 %：ID-LC/MS/MS方法测定血清Hcy的方法精密度和真实度均达到最佳水平，可用于EQA样本的定值，为监测临床实验室Hcy测定的准确性提供可靠数据。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Clinica Chimica Acta 医学-医学实验技术

CiteScore

10.10

自引率

2.00%

发文量

1268

审稿时长

23 days

期刊介绍： The Official Journal of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) Clinica Chimica Acta is a high-quality journal which publishes original Research Communications in the field of clinical chemistry and laboratory medicine, defined as the diagnostic application of chemistry, biochemistry, immunochemistry, biochemical aspects of hematology, toxicology, and molecular biology to the study of human disease in body fluids and cells. The objective of the journal is to publish novel information leading to a better understanding of biological mechanisms of human diseases, their prevention, diagnosis, and patient management. Reports of an applied clinical character are also welcome. Papers concerned with normal metabolic processes or with constituents of normal cells or body fluids, such as reports of experimental or clinical studies in animals, are only considered when they are clearly and directly relevant to human disease. Evaluation of commercial products have a low priority for publication, unless they are novel or represent a technological breakthrough. Studies dealing with effects of drugs and natural products and studies dealing with the redox status in various diseases are not within the journal''s scope. Development and evaluation of novel analytical methodologies where applicable to diagnostic clinical chemistry and laboratory medicine, including point-of-care testing, and topics on laboratory management and informatics will also be considered. Studies focused on emerging diagnostic technologies and (big) data analysis procedures including digitalization, mobile Health, and artificial Intelligence applied to Laboratory Medicine are also of interest.