Establishment of a Direct Competitive ELISA for Camel FGF21 Detection

文献类型： 外文期刊

第一作者： Yang, Yuxuan

作者： Yang, Yuxuan;Zhao, Shuqin;Gao, Yuan;Yang, Yuxuan;Yuan, Hong;Jiao, Yunjuan;Fu, Yuanfang;Bai, Xingwen;Lu, Zengjun

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关键词： camel; FGF21; prokaryotic expression; affinity chromatography purification; polyclonal antibody; direct competition ELISA

期刊名称：VETERINARY SCIENCES （ 影响因子：2.3； 五年影响因子：2.4 ）

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年卷期： 2025 年 12 卷 2 期

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收录情况： SCI

摘要： The camel is an amazing domestic species in arid and semi-arid desert regions, with multiple uses such as transportation, milk, meat, racing, and tourism. Camels can store excess fat in their humps, which enables them to survive in drought and eat less for a long period of time. Fibroblast growth factor (FGF) 21 is a key hormone that regulates metabolic pathways and energy homeostasis. However, the absence of a specific detection method for camel FGF21 impacts research on camels' metabolic regulation. This study has established a direct competition ELISA assay for detecting camel FGF21, which would provide a rapid quantitative tool for conducting research on the FGF21 factor in camels. Camels, with the ability to survive under drought and chronic hunger, developed exceptional efficient lipid reserves and energy substance metabolic characteristics. Fibroblast growth factor (FGF) 21 is a hormone that regulates important metabolic pathways and energy homeostasis. However, the absence of a specific detection method for camel FGF21 impacts research on camels' metabolic regulation. This study established a direct competition ELISA assay for detecting camel FGF21. Camel FGF21 antigen was expressed and purified through prokaryotic expression system. Polyclonal antibody was produced and purified via immunizing guinea pigs and affinity chromatography assay. Biotin-labeled FGF21 was synthesized artificially as the competitive antigen. After the determination of optimal conditions, including the working concentrations of the antibody and antigen, blocking solution, dilution buffer, and the competition reaction time, the standard curve with a typical "S" shape was generated using GraphPad Prism. The regression equation was Y = 0.1111 + (X-0.7894) x (2.162 - 0.1111)/(X-0.7894 + 15.76(-0.7894)), with the IC50 15.59 ng/mL, the limit of detection (LOD) 0.024 ng/mL, the limit of quantification (LOQ) 1.861 ng/mL, and the linear range IC20 similar to IC80 2.0 similar to 119.22 ng/mL. The verification test showed that the recovery rate ranged from 91.34% to 98.9%, and the coefficients of variation for the intra- and inter-plate both were less than 10%, indicating that the ELISA method had high accuracy, good repeatability, and high stability. In addition, this ELISA method had the potential to detect FGF21 secretion levels in other species such as mouse, human, and pig. This study provided a rapid quantitative tool for conducting research on the FGF21 factor in camels.

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