基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究

朱琳; 陈子键; 罗林; 王宇; 钟玉心; 沈玉栋; 孙远明; 徐振林

doi:10.19969/j.fxcsxb.21040903

您当前的位置：

首页 >

文章列表页 >

基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究

研究报告 | 更新时间：2023-01-04

- 基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究
- Study on Detection of Carmine by Indirect Competitive ELISA Based on Highly Specific Monoclonal Antibody
- 分析测试学报 2022年41卷第1期页码：128-135
- 作者机构：
  
  1.广东省食品质量安全重点实验室，华南农业大学食品学院，广东广州 510640
  2.广州市食品检验所，广东广州 510410
- 作者简介：
  
  徐振林，博士，教授，研究方向：食品及环境中小分子化学有害物检测技术研究与应用，E-mail： jallent@163.com
- 基金信息：
  
  十三五国家重点研发计划课题(2017YFC1601700);广东省普通高校重点研究项目(2019KJDXM002)
- DOI：10.19969/j.fxcsxb.21040903
  中图分类号：
扫描看全文
朱琳,陈子键,罗林等.基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究[J].分析测试学报,2022,41(01):128-135.

ZHU Lin,CHEN Zi-jian,LUO Lin,et al.Study on Detection of Carmine by Indirect Competitive ELISA Based on Highly Specific Monoclonal Antibody[J].Journal of Instrumental Analysis,2022,41(01):128-135.
朱琳,陈子键,罗林等.基于高特异性单克隆抗体的间接竞争ELISA检测食品中胭脂红的研究[J].分析测试学报,2022,41(01):128-135. DOI： 10.19969/j.fxcsxb.21040903.

ZHU Lin,CHEN Zi-jian,LUO Lin,et al.Study on Detection of Carmine by Indirect Competitive ELISA Based on Highly Specific Monoclonal Antibody[J].Journal of Instrumental Analysis,2022,41(01):128-135. DOI： 10.19969/j.fxcsxb.21040903.

摘要

该研究设计合成了一种胭脂红半抗原，分别采用重氮化法和戊二醛法将半抗原与载体蛋白偶联制备人工抗原，通过免疫Bal b/c小鼠及杂交瘤技术成功筛选制备了胭脂红高特异性单克隆抗体，与苋菜红、柠檬黄等结构类似物无交叉反应。基于该抗体建立了间接竞争酶联免疫分析方法用于检测食品中胭脂红残留。该方法对胭脂红的半抑制浓度（IC,50,）和检出限（IC,10,）分别为10.1 ng/mL和0.98 ng/mL，线性范围为2 ~ 50 ng/mL。将该方法应用于山楂条和雪碧中胭脂红的快速检测，样品加标回收率分别为93.0% ~ 113%和93.0% ~ 100%，相对标准偏差分别为9.7% ~ 12%和3.2% ~ 3.9%。结果与HPLC－UV方法一致，表明该方法可用于食品中痕量胭脂红的快速筛查。

Abstract

Carmine is one of the most widely used synthetic pigments in foods．However，a certain dose of carmine may result in teratogenicity，sensitization，neurotoxicity，reproductive and developmental toxicity．In order to develop a simple，rapid and accurate analytical method for screening carmine residues in food，a novel hapten against carmine was designed and synthesized in this paper. It was conjugated to lactoferrin by diazotization method to obtain immunogen and bovine serum albumin by glutaraldehyde method to obtain coating antigen．After immunizing the Bal b/c mice by immunogen，the spleen cells were fused with SP 2/0 to obtain the hybridoma screening monoclonal antibodies with high specificity to carmine．Negligible cross-reactivity with carmine analogous such as amaranth，lemon yellow，brilliant blue and sunset yellow was obtained．Based on the obtained monoclonal antibody，a simple，rapid and sensitive indirect competitive enzyme-linked immunosorbent assay（icELISA） was developed and validated. After optimizing the working conditions，the proposed icELISA showed a good sensitivity for carmine detection， with a half inhibition rate（IC,50,） of 10.1 ng/mL，a limit of detection（IC,10,） of 0.98 ng/mL and a linear range of 2－50 ng/mL．The proposed assay was then applied to detection of carmine in sprite and hawthorn strips samples．Recoveries of 93.0%－100% for sprite samples and 93.0%－113% for hawthorn strips samples were obtained with relative standard deviations of 3.2%－3.9% and 9.7%－12%，respectively，which were validated by standard high performance liquid chromatography coupled with ultraviolet-visible spectroscopy（HPLC－UV）．Results showed that the proposed icELISA was an ideal tool for the rapid screening of carmine residues in food samples.

关键词

胭脂红半抗原设计单克隆抗体间接竞争ELISA食品

Keywords

carminehapten designmonoclonal antibodyindirect competitive enzyme-linked immunosorbent assay （icELISA）food

references

GB/T 2760－2014. National Food Safety Standard－Standard for the Use of Food Additives. National Standards of the People's Republic of China（食品安全国家标准－食品添加剂使用标准．中华人民共和国国家标准）.

Golka K，Kopps S，Myslak Z W．Toxicol. Lett.，2004，151（1）：203－210.

Rundle C W，Jacob S E，Machler B C．Dermatitis，2018，29（5）：244－249.

Grant D，Gaunt I F，Carpanini F M．Food Chem. Toxicol.，1987，25（12）：913－917.

Guangdong Provincial Market Supervision Administration. 26 batches of food are unqualified.［2020－04－24］. http：//www.ipraction.gov.cn/article/gzdt/zlbg/202004/309859.htmlhttp://www.ipraction.gov.cn/article/gzdt/zlbg/202004/309859.html

Jiangxi Provincial Market Supervision Administration. 2 batches of fruit products failed to pass the random inspection.［2020－09－15］. http：//www.ipraction.gov.cn/article/gzdt/zlbg/202009/322967.htmlhttp://www.ipraction.gov.cn/article/gzdt/zlbg/202009/322967.html

Henan Provincial Market Supervision Administration. 10 batches of food are unqualified.［2020－08－27］ http：//www.ipraction.gov.cn/article/gzdt/zlbg/202008/321284.htmlhttp://www.ipraction.gov.cn/article/gzdt/zlbg/202008/321284.html

Lim H，Choi J，Song S，Kim M．Food Chem.，2014，158：521－526.

Hu Q，Sun H J，Liu L，Xiao L F，Yang Z Q，Rao S Q，Gong X J，Han J．Food Addit. Contam. A，2021. DOI：10.1080/19440049.2021.1889045http://dx.doi.org/10.1080/19440049.2021.1889045.

Xie M C，Yang J Y，Li R，Xiao Z L，Wang H，Xu Z L，Sun Y M，Shen Y D．J. Instrum. Anal. 谢美婵，杨金易，李然，肖治理，王弘，徐振林，孙远明，沈玉栋. 分析测试学报），2021，40（9）：1380－1385.

Liu Z W，Hua Q C，Wang J， Liang Z Q，Li J，Wu J H，Shen X，Lei H T，Li X M．Biosens. Bioelectron.，2020，158：112178.

Zhao Z G，Luo J Y，Fu Y W，Xu Y Y，Wang C J，Yang S H，Yang M H．J. Instrum. Anal. 赵志高，骆骄阳，付延伟，徐媛媛，王长健，杨世海，杨美华. 分析测试学报），2021，40（1）：149－158.

Xing Y．Development of Indirect Competitive ELISA for the Detection of Sunset Yellow and Ponceau 4R in Food Samples. Tianjin：Nankai University（邢http://html.publish.founderss.cn/rc-pub/api/common/picture?pictureId=37282126&type=http://html.publish.founderss.cn/rc-pub/api/common/picture?pictureId=37282129&type=http://html.publish.founderss.cn/rc-pub/api/common/picture?pictureId=37282127&type=2.799997332.88017964. 日落黄和胭脂红酶联免疫吸附测定法的建立. 天津：南开大学），2013.

Wang X．Preparation of Monoclonal Antibodies Against Ponceau 4R and Development of an ELISA Method. Yangzhou：Yangzhou University（王兴. 胭脂红单克隆抗体的制备及ELISA检测方法的建立. 扬州：扬州大学）， 2016.

Chen Z J，Liu X X，Xiao Z L，Fu H J，Huang Y P，Huang S Y，Shen Y D ，He F，Yang X X，Hammock B，Xu Z L．Ecotoxicol. Environ. Saf.， 2020， 196：110533.

Zai J J，Yi K，Xie L L，Zhu J P，Feng X T，Li Y M．Diagn. Pathol.，2018，13（1）：96.

Xu H J，Dong Y，Guo J，Jiang X M， Liu J，Xu S，Wang H Y．Toxicon，2018，156：1－6.

Wei L J，Yang Y H，Sun D Y．Food Chem.，2020. DOI：https：//doi.org/10.1016/j.foodchem.2020.127177http://dx.doi.org/https：//doi.org/10.1016/j.foodchem.2020.127177.

Li C L，Luo X S，Li Y H，Yang H J，Liang X，Wen K，Cao Y X，Li C，Wang W Y，Shi W M，Zhang S X，Yu X Z，Wang Z H．Molecules， 2019， 24（3）：443.

GB/T 5009. 35－2016. National Food Safety Standard－Determination of Synthetic Colorants in Food. National Standards of the People's Republic of China（食品安全国家标准－食品添加剂使用标准. 中华人民共和国国家标准）.

Zhang S W，Wu Z Y，Zhou K，Luo L，Xu Z L．Food Agric. Immunol.， 2019， 30（1）：594－605.

Ceballos－Alcantarilla E，Lopez－Puertollano D，Agullo C，Abad－Fuentes A，Abad－Somovilla A，Mercader J V. Analyst，2018， 143（23）：5718－5727.

Bauters T，Vandenbroucke J．J. Oncol. Pharm. Practice， 2019，25（1）：187－191.

Mahmuda A，Bande F，Kadhim Al－Zihiry K J，Abdulhaleem N，Majid R A，Hamat R A，Abdullah W O，Trop Unyah Z．J. Pharm. Res.，2017，16（3）：713－722.

Yang F，Wang H，Yang J，Yue Y P，Ke M Q，Li H Y，Wan D Y，He F R．Food Anal. Methods，2017，10（11）：3687－3693.

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

胶体金免疫层析法快速检测水产品中河豚毒素的研究

水果及环境水样中苯醚甲环唑的高灵敏免疫分析方法研究

高分辨质谱在食品农药残留检测中的研究进展

磁性碳材料的制备及其在食品农药残留检测中的应用