基于蛋白A-琼脂糖凝胶黄曲霉毒素B1免疫亲和柱的制备

doi:10.7506/spkx1002-6630-201510043

食品科学

基于蛋白A-琼脂糖凝胶黄曲霉毒素B1免疫亲和柱的制备

龚燕1，杨婷婷1，莫晓嵩2，张海涛1，黄伟2，陆廷瑾1，张东升1,*，汪海峰3

1.江苏省苏微微生物研究有限公司，江苏无锡 214063；2.江苏省粮食局粮油质量监测所，江苏南京 210011；
3.南京财经大学食品科学与工程学院，江苏南京 210011

出版日期:2015-05-25 发布日期:2015-05-08
通讯作者: 张东升
基金资助:
无锡市农业科技支撑项目（CLE02N1405）

Preparation of Immunoaffinity Column for AFB1 with Monoclonal Antibodies Immobilized on Protein A-Sepharose

GONG Yan1, YANG Tingting1, MO Xiaosong2, ZHANG Haitao1, HUANG Wei2, LU Tingjin1, ZHANG Dongsheng1,*, WANG Haifeng3

1. Jiangsu Suwei Institute of Microbiology Co. Ltd., Wuxi 214063, China;
2. Grain and Oil Quality Monitoring Bureau of Grain in Jiangsu Province, Nanjing 210011, China;
3. College of Food Science and Engineering, Nanjing University of Finance and Economics, Nanjing 210011, China

Online:2015-05-25 Published:2015-05-08
Contact: ZHANG Dongsheng

摘要/Abstract

摘要：

将初步纯化的抗黄曲霉毒素B1单克隆抗体，定向偶联于蛋白A-琼脂糖凝胶，制备成免疫亲和层析柱。结果显示，辛酸-饱和硫酸铵纯化后的单抗质量浓度为17.5 mg/mL，纯度为45.1%，抗体亚型为IgG1。取纯化后抗体48 μL与1 mL蛋白A凝胶偶联，偶联率达98.1%，高效液相色谱测定其平均相对柱容量为105 ng/0.125 mL凝胶、柱空白为0，不同样品（玉米粉、面粉、花生油、酱油、醋等）平均加标回收率在80%以上。该亲和柱在2～8 ℃保存12 个月后，相对柱容量仍可达72 ng/0.125 mL凝胶。该法制备的亲和柱相对柱容量大，稳定性好，可用于样品中黄曲霉毒素B1分离净化的要求。

关键词: 黄曲霉毒素B1, 蛋白A-琼脂糖凝胶, 定向偶联, 免疫亲和柱

Abstract:

The immunoaffinity column (IAC) for aflatoxin B1 (AFB1) was prepared by directed coupling of protein
A-sepharose with the anti-AFB1 monoclonal antibody which was preliminarily purified by caprylic acid-ammonium sulfate
method. The results showed that the concentration, purity and antibody subtype of the purified monoclonal antibody were
17.5 mg/mL, 45.1% and IgG1, respectively. When coupling 48 μL of purified antibody with protein A-sepharose, and
the coupling rate was 98.1%. The average relative column capacity and column blank were 105 ng/0.125 mL gel and 0,
respectively. The average recoveries of standard addition in different samples (corn flour, wheat flour, peanut oil, soy sauce)
were all above 80%. After being stored at 2–8 ℃ for 12 months, the relative capacity of the IAC was 72 ng/0.125 mL gel.
These results show that the anti-AFB1 affinity column prepared by this method has large relative capacity and good stability
which meet the requirement for separation and purification of AFB1 from real samples.

Key words: aflatoxin B1, protein A-sepharose, directed coupling, immunoaffinity column

中图分类号:

O657

龚燕1，杨婷婷1，莫晓嵩2，张海涛1，黄伟2，陆廷瑾1，张东升1,*，汪海峰3. 基于蛋白A-琼脂糖凝胶黄曲霉毒素B1免疫亲和柱的制备[J]. 食品科学, doi: 10.7506/spkx1002-6630-201510043.

GONG Yan1, YANG Tingting1, MO Xiaosong2, ZHANG Haitao1, HUANG Wei2, LU Tingjin1, ZHANG Dongsheng1,*, WANG Haifeng3. Preparation of Immunoaffinity Column for AFB1 with Monoclonal Antibodies Immobilized on Protein A-Sepharose[J]. FOOD SCIENCE, doi: 10.7506/spkx1002-6630-201510043.

[1]	王邹璐琪，李立煌，李丹阳，艾超超，任磊，孙本强. 构建下转换荧光-适配体的免疫层析试纸条用于快速检测黄曲霉毒素B1[J]. 食品科学, 2021, 42(12): 295-301.
[2]	张居典，刘亚文，王喜泉，徐珒昭，徐境含，武明月，许晓曦. 鼠李糖乳杆菌对大鼠黄曲霉毒素B1促排效果的评价[J]. 食品科学, 2020, 41(9): 119-125.
[3]	张敏，张先舟，李聪，高浩，刘健慧，田益玲，马雯，李英军，檀建新. 黄曲霉毒素B1核酸适配体筛选和检测方法的建立[J]. 食品科学, 2020, 41(24): 295-303.
[4]	吴宇，叶金，李丽，李森，轩志宏，崔华，刘洪美，王松雪，马海华，张峰，何建洪. 免疫亲和柱全自动净化-超高效液相色谱-串联质谱-全碳标记内标法精确测定粮油中玉米赤霉烯酮[J]. 食品科学, 2020, 41(24): 267-272.
[5]	郑婷婷, 刘玉兰, 王戬东, 裴娅晓. 淡碱蒸胚对玉米胚及其毛油中真菌毒素的降解消除作用[J]. 食品科学, 2020, 41(13): 8-13.
[6]	张威，张文中，郭平，胡重怡，吕小丽. 食用油中黄曲霉毒素B1快速检测试纸的评价[J]. 食品科学, 2020, 41(12): 326-331.
[7]	刘英丽，毛慧佳，杨梓妍，万真，王静，孙宝国. 重组漆酶降解黄曲霉毒素B1分子对接分析及产物结构解析[J]. 食品科学, 2019, 40(24): 119-127.
[8]	付凌萌，吴逸，王菁，魏婕，王少康，孙桂菊. 木犀草素与叶酸对黄曲霉毒素B1致食管上皮细胞毒性及MTHFR基因高甲基化的影响[J]. 食品科学, 2019, 40(23): 142-150.
[9]	杨威，魏学鼎，雷芬芬，胡传荣，何东平，罗质，郑竟成. 4 种方法脱除花生油中黄曲霉毒素B1的研究[J]. 食品科学, 2019, 40(22): 339-346.
[10]	赵萌，高婧，褚华硕，梁志宏. 黄曲霉毒素B1的分子致毒机理及其微生物脱毒研究进展[J]. 食品科学, 2019, 40(11): 235-245.
[11]	张牧臣，郑楠，王加启. 食品中黄曲霉毒素B1污染研究进展[J]. 食品科学, 2018, 39(7): 312-320.
[12]	王向阳，顾双，杨玲. 橘霉素免疫牛蛙肌肉蛋白柱的制备及其吸附橘霉素作用分析[J]. 食品科学, 2018, 39(22): 159-165.
[13]	王强，王旭峰，赵东豪，黎智广，蔡楠，关婉琪，黄珂，李刘冬. 免疫亲和柱净化-UPLC-MS/MS测定鱼虾肉中的3 种氨基糖苷类抗生素[J]. 食品科学, 2018, 39(20): 326-331.
[14]	于慧春，娄楠，殷勇，刘云宏. 基于高光谱技术及SPXY和SPA的玉米毒素检测模型建立[J]. 食品科学, 2018, 39(16): 328-335.
[15]	赵春霞，王轶，程薇，吕育财，龚大春，郭鹏. 复合菌系降解黄曲霉毒素B1的效果及组成多样性研究[J]. 食品科学, 2017, 38(9): 106-112.

基于蛋白A-琼脂糖凝胶黄曲霉毒素B1免疫亲和柱的制备

Preparation of Immunoaffinity Column for AFB1 with Monoclonal Antibodies Immobilized on Protein A-Sepharose

RichHTML

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价