Immunochemical determination of oxytetracycline in fish: Comparison between enzymatic and time-resolved fluorometric assays
Immunochemical determination of oxytetracycline in fish: Comparison between enzymatic and time-resolved fluorometric assays
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DOI：
10.1016/j.aca.2009.12.044
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发表时间：
2010-03-10
影响因子：
6.2
通讯作者：
Espinos, Francisco
中科院分区：
文献类型：
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作者：
Chafer-Pericas, Consuelo;Maquieira, Angel;Espinos, Francisco
建立了间接竞争酶联免疫吸附试验（ELISA）-辣根过氧化物酶（HRP）检测法，用于检测黄鲷（Sparus aurata）样品中土霉素（OTC）的含量。结果进行了比较，通过时间分辨荧光免疫测定法（TR-FIA）使用的第二抗体与粪卟啉铂（II）（PtCP）作为标记物。在鱼提取物中获得的检测限为16和0.08 μ g kg（-1）的光度和荧光检测，分别为，因此，他们是适合于鱼的质量控制，根据最大残留水平由欧盟制定的。一个提取程序，使用甲醇：水70：30（v/v）+1毫升EDTA 0.1 M，和不同的清洁程序的基础油固相萃取（C-18，聚合物反相，SCX，Si）进行了测定。采用不含四环素的鱼肉提取物标准曲线定量，克服了基质效应，同时将优化的OTC ELISA方法用于四环素和金霉素残留的检测，结果良好。因此，所开发的免疫测定法可以被认为是水产养殖抗生素治疗中最常用的四环素类抗生素的通用测定法。为了确认所开发的免疫测定法作为半定量方法的实用性，对从不同超市获得的鱼样品进行了分析。结果与参考HPLC方法所得结果相关性良好。(C)2010爱思唯尔有限公司版权所有。
An indirect competitive enzyme-linked immunosorbent assay (ELISA) with photometric detection of horseradish peroxidase (HRP) activity, was developed in plate to detect oxytetracycline (OTC) in Gilthead sea bream (Sparus aurata) samples. The results were compared to those obtained by time-resolved fluoroimmunoassay (TR-FIA) using a secondary antibody with coproporphyrin of platinum (II) (PtCP) as marker. The limits of detection obtained in fish extract were 16 and 0.08 mu g kg(-1) for photometric and fluorometric detections, respectively; therefore, they were suitable for fish quality control according to the maximum residue level established by the European Union.An extraction procedure using methanol:water 70:30 (v/v) + 1 mL EDTA 0.1 M, and different clean-up procedures based oil solid-phase extraction (C-18, polymeric reversed phase, SCX, Si) was assayed. The matrix effects were overcome by means of an average tetracycline-free fish extract calibration curve used for quantification.The OTC optimized ELISA can also be applied to determine tetracycline and chlortetracycline residues with good results. Thus, the developed immunoassay could be considered as a generic assay for the most used tetracyclines in aquaculture antibiotic treatments.In order to confirm the utility of the developed immunoassay as a semi-quantitative methodology, fish samples obtained from different supermarkets were analyzed. Results correlate well with those obtained with a reference HPLC method. (C) 2010 Elsevier B.V. All rights reserved.