Rapid determination of six fluoroquinolones in aquatic product by direct competitive enzyme-linked immunosorbent assay
-
摘要: 采用过碘酸钠法将抗环丙沙星单克隆抗体(MAb-CIP)与辣根过氧化物酶(HRP)偶联制备酶标抗体MAbCIP-HRP,建立了检测水产品中6种氟喹诺酮类药物残留的直接竞争酶联免疫吸附分析方法(dcELISA),考察了包被原浓度、竞争反应时间和有机溶剂等因素对方法灵敏度的影响。结果表明:在优化的反应条件下,所建立的dcELISA针对环丙沙星、恩诺沙星、诺氟沙星、培氟沙星、沙拉沙星和双氟沙星6种氟喹诺酮类药物的检测限(LOD)均不超过0.5ng/mL,线性范围(IC20IC80)在1.012.1ng/mL之间;对虾、鳗鲡和鲫鱼三种水产样品中添加5.0、10.0和20μg/kg时,加标回收率为70.4%104.1%,相对标准偏差为5.0%14.7%;本方法可用于水产品中氟喹诺酮类(FQs)药物多残留的快速测定。Abstract: In this study, a direct competitive enzyme-linked immunosorbent assay (dcELISA) was developed for rapid determination six fluoroquinolones in aquatic product. Horseradish peroxidase-labelled ciprofloxacin monoclonal antibody ( MAb-CIP-HRP) was synthesized by NaIO4method. The effect of coating antigen concentration, competitive reaction time and organic solvents on the sensitivity of dcELISA were investigated.Under the optimized assay conditions, the dcELISA showed the limit of detection below 0.5ng/mL for ciprofloxacin, enrofloxacin, norfloxacin, pefloxacin, sarafloxacin and difloxacin, and the linear range (IC20IC80) of 1.0 to 12.1ng/mL.Recoveries from spiked shrimp, eel and crucian at levels of 5.0, 10 and 20μg/kg were in the range of 70.4%104.1%, with relative standard deviation ranging from 5.0% to 14.7%. Results indicated that the established dcELISA was suitable for multi-analytes rapid determination of fluoroquinolones residues in aquatic product.
-
Keywords:
- fluoroquinolone /
- antibody /
- direct competitive ELISA /
- aquatic product
-
[1] Wang Z, Zhu Y, Ding S, et al.Development of a monoclonal antibody-based broad-specificity ELISA for fluoroquinolone antibiotics in foods and molecular modeling studies of crossreactive compounds[J].Anal Chem, 2007, 79 (12) :4471-4483.
[2] Moema D, Nindi M M, Dube S.Development of a dispersive liquid-liquid microextraction method for the determination of fluoroquinolones in chicken liver by high performance liquid chromatography[J].Anal Chim Acta, 2012, 730:80-86.
[3] 王战辉, 沈建忠, 张素霞.喹诺酮类药物抗体制备研究进展及策略分析[J].中国农业科学, 2008, 41 (10) :3311-3317. [4] Lee S, Kim B, Kim J.Development of isotope dilution-liquid chromatography tandem mass spectrometry for the accurate determination of fluoroquinolones in animal meat products:optimization of chromatographic separation for eliminating matrix effects on isotope ratio measurements[J].J Chromatogr A, 2013, 1277:35-41.
[5] Leivo J, Chappuis C, Lamminmaki U, et al.Engineering of a broad-specificity antibody:detection of eight fluoroquinolone antibiotics simultaneously[J].Anal Biochem, 2011, 409 (1) :14-21.
[6] Herrera-Herrera A V, Hernandez-Borges J, RodriguezDelgado M A.Fluoroquinolone antibiotic determination in bovine, ovine and caprine milk using solid-phase extraction and high-performance liquid chromatography-fluorescence detection with ionic liquids as mobile phase additives[J].J Chromatogr A, 2009, 1216 (43) :7281-7287.
[7] Kantiani L, Farre M, Barcelo D.Rapid residue analysis of fluoroquinolones in raw bovine milk by online solid phase extraction followed by liquid chromatography coupled to tandem mass spectrometry[J].J Chromatogr A, 2011, 1218 (50) :9019-9027.
[8] Hernandez M, Aguilar C, Borrull F, et al.Determination of ciprofloxacin, enrofloxacin and flumequine in pig plasma samples by capillary isotachophoresis-capillary zone electrophoresis[J].J Chromatogr B Analyt Technol Biomed Life Sci, 2002, 772 (1) :163-172.
[9] Li J, Liu J, Zhang H C, et al.Broad specificity indirect competitive immunoassay for determination of nitrofurans in animal feeds[J].Anal Chim Acta, 2010, 678 (1) :1-6.
[10] Shen Y D, Deng X F, Xu Z L, et al.Simultaneous determination of malachite green, brilliant green and crystal violet in grass carp tissues by a broad-specificity indirect competitive enzyme-linked immunosorbent assay[J].Anal Chim Acta, 2011, 707 (1-2) :148-154.
[11] Emmanuella Uwamahoro, 吕书为, 唐秋实, 等.基于洛美沙星免疫原的喹诺酮药物广谱特异性抗体制备的研究[J].食品工业科技, 2012, 33 (20) :156-159. [12] 杨正涛, 张乃生, 魏东.氟喹诺酮类药物多组份残留的酶免疫分析法[J].中国兽医学报, 2008, 28 (9) :1057-1060. [13] Lei H T, Shen Y D, Song L J, et al.Hapten synthesis and antibody production for the development of a melamine immunoassay[J].Anal Chim Acta, 2010, 665 (1) :84-90.
[14] Xu Z L, Deng H, Deng X, 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.
[15] 杨星星, 刘细霞, 王弘, 等.细交链孢菌酮酸酶联免疫吸附分析方法研究[J].分析化学, 2012, 40 (9) :1347-1352. [16] Esteve-Turrillas F A, Abad-Fuentes A, Mercader J V.Determination of fenhexamid residues in grape must, kiwifruit, and strawberry samples by enzyme-linked immunosorbent assay[J].Food Chemistry, 2011, 124 (4) :1727-1733.
计量
- 文章访问数: 122
- HTML全文浏览量: 10
- PDF下载量: 194