中国兽医科学2025，Vol.55Issue(6)：764-769,6.DOI:10.16656/j.issn.1673-4696.2025.0087
牛C-反应蛋白夹心ELISA定量检测方法的建立与应用
Establishment and application of a sandwich ELISA method for quantitative detection of bovine C-reactive protein
摘要
Abstract
This study aimed to develop monoclonal antibodies against bovine C-reactive protein(CRP)and establish a double-antibody sandwich ELISA method for quantitative detection of bovine cytokines,offering a tool for rapid infection diagnosis.Bovine CRP was expressed using prokaryotic and 293F eukaryotic expression systems,followed by recombinant protein purification through affinity chromatography.Monoclonal antibodies were generated via immunization of mice and lymphocyte hybridoma technology,and a sandwich ELISA was developed using checkerboard analysis of antibody pairs.The method was validated with clinical samples.SDS-PAGE result confirmed that CRP was purified.Five monoclonal antibody cell lines were established,with 3H4 and 2G4-HRP pairs demonstrating optimal detection performance.Clinical sample analysis showed significantly elevated CRP levels in serum of cattle in the early stages of tuberculosis compared to healthy cattle.This suggests that the produced monoclonal antibodies can identify natural bovine CRP and effectively measure CRP levels in plasma samples from tuberculosis-infected cattle,with incubation period cattle showing significantly higher CRP mass concentrations than those in the active period cattle.In conclusion,the developed sandwich ELISA method reliably diagnoses bovine tuberculosis,and provides a foundation for evaluating immune status and diagnosing infectious diseases in cattle.关键词
C-反应蛋白/单克隆抗体/ELISA/牛结核病Key words
C-reactive protein/monoclonal antibodies/ELISA/bovine tuberculosis分类
农业科技引用本文复制引用
靳家鑫,杨蕊,张泽,时文健,张广智,丁家波,鑫婷..牛C-反应蛋白夹心ELISA定量检测方法的建立与应用[J].中国兽医科学,2025,55(6):764-769,6.基金项目
北京市自然科学基金项目(6164039) （6164039）
宁夏回族自治区重点研发计划项目(2024BBF02014) （2024BBF02014）
中央级公益性科研院所基本科研业务费专项(2022-YWF-ZYSQ-01) （2022-YWF-ZYSQ-01）
