Enzyme-Linked Immunosorbent Assay for Pork Determination in Raw and Heated Meats: Combination of Monoclonal Antibodies to Denatured Porcine Myoglobin and Sodium Dodecyl Sulfate Extraction
Enzyme-Linked Immunosorbent Assay for Pork Determination in Raw and Heated Meats: Combination of Monoclonal Antibodies to Denatured Porcine Myoglobin and Sodium Dodecyl Sulfate Extraction
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用于生肉和加热肉中猪肉测定的酶联免疫吸附测定：变性猪肌红蛋白单克隆抗体与十二烷基硫酸钠提取的组合
DOI：
10.1021/acsfoodscitech.1c00372
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Narita Hiroshi
中科院分区：
文献类型：
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作者：
Yamasaki Tomomi;Hirakawa Yuki;Momma Keiko;Yamaguchi Yukie M.;Kotoura Satoshi;Miyake Shiro;Narita Hiroshi
已经开发了几种用于监测猪肉污染的免疫测定法，但它们在准确检测方面存在局限性，因为它们受到蛋白质变性和烹饪导致的提取效率损失的影响。本文建立了一种结合十二烷基硫酸钠（SDS）提取方法的双抗体夹心酶联免疫吸附试验（s-ELISA）检测生肉和加热肉制品中猪肉残留的方法。为了建立单克隆抗体（mAb），将SDS变性的猪肌红蛋白（Mb）和具有猪Mb氨基酸序列的合成肽用于免疫。该方法可定量检测猪肌红蛋白，与牛肉、鸡肉、羊肉、山羊肉无交叉反应。猪肌红蛋白用于s-ELISA的50%最大有效浓度为90 ng/mL，加标回收率为94- 158%。该s-ELISA检测到1%（w/w）的猪肉与生的和加热的牛肉混合。对于测定不同部位的生猪肉，包括脂肪腹部的肌红蛋白，结果相关性良好，通过高效液相色谱法得到的。本研究建立的s-ELISA方法可用于肉制品的鉴别和防止猪肉产品的标签错误。
Several immunoassays for monitoring pork contamination have been developed, but they have limitations in accurate detection because they have been affected by protein denaturation and loss of extraction efficiency due to cooking. In this study, a sandwich enzyme-linked immunosorbent assay (s-ELISA) combined with an extraction method using sodium dodecyl sulfate (SDS) was developed for pork determination in raw and heated meats. To establish monoclonal antibodies (mAbs), SDS-denatured porcine myoglobin (Mb) and synthetic peptides with amino acid sequences of porcine Mb were used for immunization. The s-ELISA quantitatively detected the porcine Mb without cross-reaction to beef and chicken Mbs and lamb and goat meats. The 50% maximal effective concentration of porcine Mb for s-ELISA was 90 ng/mL, and the recoveries of porcine Mb spiked into raw and heated beef and chicken were 94–158%. This s-ELISA detected 1% (w/w) pork mixed with raw and heated beef. For the determination of Mbs in various parts of raw pork, including fatty bellies, the results correlated well with those obtained by high-performance liquid chromatography. Our s-ELISA could be available for authentication of meat products and prevention of mislabeling in pork products.