文献类型：期刊文献

英文题名：Development of an indirect competitive ELISA combining optimized extraction and heat-stable epitope targeting for crustacean allergen detection

作者：Zhu, Wenye[1,2];Fu, Qi[1];Wang, Hao[1];Zhang, Ziye[1];Liu, Zhiyu[3];Xu, Min[3];Lin, Hong[1];Deng, Junming[2];Tan, Beiping[2];Li, Zhenxing[1]

机构：[1]Ocean Univ China, Coll Food Sci & Engn, State Key Lab Marine Food Proc & Safety Control, Huangdao Campus, 1299, Sansha Rd, Qingdao 266000, Shandong, Peoples R China;[2]Guangdong Ocean Univ, Coll Fisheries, 1, Haida Rd, Zhanjiang 524088, Guangdong, Peoples R China;[3]Fisheries Res Inst Fujian, 7, Haishan Rd, Xiamen 361013, Fujian, Peoples R China

年份：2027

卷号：193

外文期刊名：FOOD CONTROL

收录：SCI-EXPANDED(收录号：WOS:001876039100001)、、WOS

基金：This research was supported by the National Natural Science Foun-dation of China (32372439) , the National Key Research and Develop-ment Program of China (2023YFD2401504) , and the China Postdoctoral Science Foundation (2026M793263) .

语种：英文

外文关键词：Crustacean allergen detection; Sarcoplasmic calcium-binding protein; Extraction buffer optimization; Heat-stable epitope; Specific affinity purification; Indirect competitive ELISA

外文摘要：Sarcoplasmic calcium-binding protein (SCP) has been proposed as a target to resolve false positives caused by cross-reactivity in crustacean allergen immunodetection. Whereas decreased detection signals induced by food thermal processing remain a challenge. To overcome this issue, the allergen extraction buffer was optimized, and the stable epitope of SCP (P7, AQFISNPDEKCNAVYL) was selected as the detection target to establish an indirect competitive ELISA (icELISA). The 50 mM carbonate buffered saline supplemented with 1% Tween 20 was identified as the optimal solution, since SCP extraction efficiency was improved by 35%-122% and no obvious interference was observed on SCP icELISA. P7-KLH antisera exhibited stable recognition toward SCP before and after autoclaved treatment, and its cross-reactivity with KLH and mollusks was eliminated by SCP affinity purification. Based on the P7-specific antibody, the P7 icELISA was developed and evaluated. Despite limited sensitivity (analytical limit of quantification: 1.06 mu g/mL SCP), the assay showed a working range of 0.1-100 mu g/ mL, SCP-spike recoveries of 84.6%-109.5%, and precision CVs of 3.7%-12.1%. No cross-reactivity was observed among the tested non-crustacean species, and detected values in selected processed foods were 11.5-50.0-fold higher than those obtained with the SCP icELISA. These results indicate the potential of heat-stable epitope targeting to reduce processing-induced signal loss in SCP immunodetection.