[1]赵扬,武爱华,张霄,等.有机磷农药广谱抗原表位多肽的制备及应用[J].江苏农业学报,2017,(02):444-449.[doi:doi:10.3969/j.issn.1000-4440.2017.02.032]
　ZHAO Yang,WU Ai-hua,ZHANG Xiao,et al.Preparation and application of broad spectrum antigen epitope peptide against organophosphorus pesticides[J].,2017,(02):444-449.[doi:doi:10.3969/j.issn.1000-4440.2017.02.032]
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有机磷农药广谱抗原表位多肽的制备及应用()
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江苏农业学报[ISSN:1006-6977/CN:61-1281/TN]

卷:
期数:
2017年02期
页码:
444-449
栏目:
加工贮藏·质量安全
出版日期:
2017-04-30

文章信息/Info

Title:
Preparation and application of broad spectrum antigen epitope peptide against organophosphorus pesticides
作者:
赵扬12; 武爱华2; 张霄2; 莫海珍1; 张存政2; 刘贤金2; 胡梁斌1
（1.河南科技学院食品学院，河南新乡453000；2.江苏省农业科学院食品质量安全与检测研究所，江苏南京210014）
Author(s):
ZHAO Yang12; WU Ai-hua2; ZHANG Xiao2; MO Hai-zhen1; ZHANG Cun-zheng2; LIU Xian-jin2; HU Liang-bin1
(1.School of Food Science,Henan Institute of Science and Technology, Xinxiang 453000, China;2.Institute of Food Quality Safety and Detection Research, Jiangsu Academy of Agricultural Sciences，Nanjing 210014, China)
关键词:
有机磷类农药; 噬菌体展示十二肽库; 模拟抗原表位多肽; 间接竞争ELISA
Keywords:
organophosphates pesticide; phage displayed random 12-mer peptide library; antigen epitope mimicking peptide; indirect competitive ELISA
分类号:
X836
DOI:
doi:10.3969/j.issn.1000-4440.2017.02.032
文献标志码:
A
摘要:
利用噬菌体展示技术，以广谱型单克隆抗体筛选十二肽噬菌体展示库，获得能够模拟有机磷类农药抗原表位的十二肽，建立了一种间接竞争ELISA方法用于检测多种有机磷类农药。结果表明，获得的特异性多肽氨基酸序列为HPAPDHSSQSHQ，呈现出较好的抗原性、亲水性及表面可及性，可广谱模拟有机磷农药抗原表位。所建立的间接ELISA方法对有机磷农药通用结构半抗原（BPB）的IC50和IC10分别为1.776 μg/ml及0.358 μg/ml，与对硫磷、蝇毒磷、乙拌磷、辛硫磷、喹硫磷和三唑磷的交叉反应率分别为58.74%、44.93%、52.03%、43.98%、36.82%和4.87%。建立的检测方法具有绿色、广谱、高效的特点，是一种新型有机磷类农药累积毒性检测技术。
Abstract:
Using phage display technology, polypeptides mimicking organophosphorus pesticides antigen epitope were obtained by broad anti-organophosphates monoclonal antibody screening. Thereby,an indirect competitive ELISA method for the multi-determination of BPB and organophosphates pesticides was established using polypeptide as immobilized antigen. The obtained sequence of polypeptide is HPAPDHSSQSHQ, featured by good antigenicity, hydrophilicity and surface accessibility. The IC50 and IC10 for the hapten BPB detected by the indirect ELISA were 1.776 μg/ml and 0.358 μg/ml, respectively. The cross-reactivities with parathion, coumaphos, disulfoton, phoxim, quinalphos and triazophos were 58.74%, 44.93%, 52.03%, 43.98%, 36.82% and 4.87%. As a green, high-throughput technology, the broad specific immunoassay provide a new technical route for the accumulative toxicity determination of organophosphorus pesticides.

参考文献/References:

[1]贺锡雯，吕京. 有机磷类和拟除虫菊酯类农药的神经毒作用机理[J].中国药理学与毒理学杂志, 1997，11(2): 89-90.
[2]林春绵,胡晓燕，张安平. 酶抑制法快速检测有机磷农药残留的研究进展[J].浙江工业大学学报, 2009，37(4): 386-391.
[3]BERIJANI S, SADIGH M, POURNAMDARI E. Homogeneous liquid-liquid microextraction for determination of organophosphorus pesticides in environmental water samples prior to gas chromatography-flame photometric detection[J]. Journal of Chromatographic Science, 2016, 54:1061-1067.
[4]SEEBUNRUENG K, SANTALADCHAIYAKIT Y, SRIJARANAI S. Vortex-assisted low density solvent based demulsified dispersive liquid-liquid microextraction and high-performance liquid chromatography for the determination of organophosphorus pesticides in water samples[J]. Chemosphere, 2014, 103: 51-58.
[5]XU Z L, DENG H, DENG X F, et al. Monitoring of organophosphorus pesticides in vegetables using monoclonal antibody-based direct competitive ELISA followed by HPLC-MS/MS[J]. Food Chemistry, 2012, 131(4): 1569-1576.
[6]WANG H, LI G H, CHEN T, et al. Immunoassay for multiple residues of organophosphorus pesticides using broad spectrum of monoclonal antibodies[J]. Journal of Food Safety and Quality, 2015, 6(11): 4399-4408.
[7]HUA X, WANG L, LI G, et al. Multi-analyte enzyme-linked immunosorbent assay for organophosphorus pesticides and neonicotinoid insecticides using a bispecific monoclonal antibody[J]. Analytical Methods, 2013, 5(6): 1556-1563.
[8]张小兵,邸禄芹,吴萌,等. 单克隆抗体与多克隆抗体配对ELISA方法比较[J].生物技术通报, 2009(11): 125-129.
[9]武爱华,张霄,刘媛,等. Cry2 Aa毒素结合十二肽的筛选与鉴定[J].江苏农业学报, 2015(4): 929-941.
[10]INABA J, NAKAMURA S, SHIMIZU K, et al. Anti-metatype peptides, a molecular tool with high sensitivity and specificity to monitor small ligands[J]. Analytical Biochemistry, 2009, 388(1): 63-70.
[11]BRIGATI J R, PETRENKO V A. Thermostability of landscape phage probes[J]. Analytical and Bioanalytical Chemistry, 2005, 382(6): 1346-1350.
[12]SCHWIND P, KRAMER H, KREMSER A, et al.Subtilisin removes the surface layer of the phage fd coat[J]. European Journal of Biochemistry, 1992,210(2): 431-436.
[13]JOHNSON H R, HOOKER J M, FRANCIS M B, et al. Solubilization and stabilization of bacteriophage MS2 in organic solvents[J]. Biotechnology and Bioengineering, 2007, 97(2): 224-234.
[14]贺江.噬菌体展示技术在食品科学领域的应用进展[J].中国农学通报, 2014，30(24): 303-309.
[15]王妍入. 黄曲霉毒素替代物绿色免疫分析研究[D]. 北京：中国农业科学院，2014.
[16]杨威,李小兵,刘国文，等. 几种方法纯化小鼠腹水IgG2b类单抗的比较[J].畜牧与兽医, 2011(1): 48-50.
[17]张霄,纪炜,王耘，等. 间接ELISA法检测苏云金芽孢杆菌(Bt)Cry1Ab毒蛋白抗体方法的建立[J].江苏农业学报, 2010，26(4): 894-896.
[18]WRIGHTON N C, FARRELL F X, CHANG R, et al. Small peptides as potent mimetics of the protein hormone erythropoietin[J]. Science, 1996, 273(5274): 458-463.
[19]ROSELYNE B, CAROLINE D, BERNARD M, et al. Identification of a peptide blocking vascular endothelial growth factor (VEGF)-mediated angiogenesis[J]. EMBO Journal, 2000, 19(7): 1525-1533.
[20]华修德. 有机磷农药单残留及多残留免疫分析方法的建立与应用[D].南京：南京农业大学，2012.

备注/Memo

备注/Memo:
收稿日期：2016-05-30 基金项目：江苏省自然科学基金项目（BK20131333）；江苏省科技支撑计划项目（BE2014722） 作者简介：赵扬（1989-），男，河南安阳人，硕士，主要从事农产品质量与安全研究。（E-mail）13913979573@163.com 通讯作者：胡梁斌，（E-mail）hulb973@163.com
更新日期/Last Update: 2017-05-02