基于电化学发光免疫分析技术定量检测人血清中司美格鲁肽的含量

摘要/Abstract

摘要： 目的：建立一种定量检测人血清中司美格鲁肽含量的电化学发光免疫分析方法，为该多肽类药物的临床定量测定提供新的检测手段。方法：通过小鼠免疫及杂交瘤细胞技术制备抗司美格鲁肽的单克隆抗体，并经多轮筛选获得最优抗体对作为本分析方法的关键试剂。利用捕获抗体、司美格鲁肽待测样品和生物素标记的侦测抗体形成夹心结构作为分析原理，通过电化学发光信号强度与待测物浓度的线性关系实现定量检测。根据方法的用途确定灵敏度和特异性等关键参数，并结合生物分析指导原则对该分析方法进行一系列方法学验证工作。结果：本研究所建立的分析方法灵敏度满足临床检测需求（2.0 ng·mL-1），标准曲线定量范围为2.0~240.0 ng·mL-1；批内及批间精密度%CV≤17.1%，批内及批间的准确度|%RE|≤17.3%，均满足接受标准；该方法对司美格鲁肽具有高度特异性，可耐受重组人GLP-1高达200.0 ng·mL-1；样本稀释在200倍以内稀释线性良好，并在司美格鲁肽浓度高达2 400.0 ng·mL-1时未观察到钩状效应；选择性和稳定性等参数的验证结果均在可接受标准之内。结论：本研究创新性地建立了基于电化学发光免疫分析方法定量检测人血清中司美格鲁肽的含量，方法开发始于单克隆抗体试剂的制备，直至电化学发光检测法的建立和验证。该方法具有高灵敏度、高特异性等特点，为司美格鲁肽的临床药代动力学研究提供了一种新的检测手段。

关键词: font-size:medium, ">司美格鲁肽；电化学发光免疫分析；方法开发；生物分析

Abstract: Objective： This study aimed to establish an electrochemiluminescence immunoassay for the quantitative determination of semaglutide in human serum， providing a novel analytical approach for the clinical quantification of this peptide-based drug. Methods： Anti-semaglutide monoclonal antibodies were prepared by mouse immunization and hybridoma technology， and the resulted monoclonal antibody reagents were screened to obtain an optimal pair for this analytical method. The assay is based on the formation of a sandwich structure from the capture antibody， semaglutide， and the biotin-labeled detection antibody. Quantification is achieved by correlating the intensity of electrochemiluminescence signals with the concentration of the semaglutide. Key parameters such as assay sensitivity and specificity were established according to the intended use of the method， and validated per bioanalytical method validation guidance. Results： The assay sensitivity was determined to be 2.0 ng·mL-1 for clinical bioanalysis of semaglutide with a quantitative range of 2.0 to 240.0 ng·mL-1. The intra- and inter-assay precision (%CV) were both ≤17.1%， and intra- and inter-assay accuracy |%RE| were ≤17.3%， all meeting the pre-set requirement. Additionally， the method exhibited adequate specificity， as recombinant human GLP-1 up to 200.0 ng·mL-1 did not interfere with semaglutide quantification. The assay displayed good dilution linearity with dilution factors up to 200， and no hook effect was observed at an analyte concentration up to 2 400.0 ng·mL-1. Other validation parameters such as selectivity and sample stability were all within acceptable standards. Conclusion： In this study， an innovative electrochemiluminescence-based immunoassay was developed and validated for quantitative determination of semaglutide in human serum. Validation data demonstrate adequate sensitivity and specificity. As a novel method， it is suitable for pharmacokinetic study of semaglutide in a clinical trial.

Key words: font-size:medium, ">Semaglutide； Electrochemiluminescence immunoassay； Method development； Biologlical analysis

中图分类号:

罗逸帆1, 2, 张贺峰2, 3, 梁宇2, 3, 邹灵龙1, 2, 3. 基于电化学发光免疫分析技术定量检测人血清中司美格鲁肽的含量[J]. 中国药物评价, 2025, 42(1): 42-47.

LUO Yifan1, 2, ZHANG Hefeng2, 3, LIANG Yu2, 3, ZOU Linglong1, 2, 3. An Electrochemiluminescence Immunoassay for Quantitative Determination of Semaglutide in Human Serum[J]. CHINESE JOURNAL OF DRUG EVALUATION, 2025, 42(1): 42-47.

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