Abstract
By combining a chemiluminescence immunoassay with microfluidic chip technology, a simple assay was established for the sensitive detection of cloxacillin in poultry samples. The chip used in this approach was composed of an upper microchannel layer and a lower flat base layer. Each of the formed microchannels has a volume of 18 μL, thus enabling a considerable reduction of reagent consumption as well as sample size needed for analysis by the indirect competitive immunoassay. To obtain labeled antigens for coating, the active ester method was employed to couple cloxacillin to glucose oxidase (GlcOx). The immobilized GlcOx-cloxacillin conjugate on the base layer competed with residues of cloxacillin in samples for binding sites of the monoclonal antibodies (mAb) against cloxacillin. Horseradish peroxidase-labeled anti-mouse IgG antibodies and chemiluminescent substrate were used for signal generation. Under optimized conditions, the IC50 and the limit of detection (LOD) of the assay were at 96.5 ± 9.37 and 0.92 ± 0.07 ng/mL, respectively, and thus well below the maximum residue level of 300 ng/mL set by the European Commission. Recoveries for spiked chicken and duck samples were in the range from 94 to 118 % with relative standard deviations lower than 11 %. The results demonstrate that the microfluidic chip-based immunoassay can be used as a rapid and reliable platform for determination of cloxacillin in poultry.
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References
Awad YM et al (2014) Veterinary antibiotics contamination in water, sediment, and soil near a swine manure composting facility. Environ Earth Sci 71:1433–1440
Babington R, Matas S, Marco MP, Galve R (2012) Current bioanalytical methods for detection of penicillins. Anal Bioanal Chem 403:1549–1566
Berendsen BJA, Gerritsen HW, Wegh RS, Lameris S, van Sebille R, Stolker AAM, Nielen MWF (2013) Comprehensive analysis of -lactam antibiotics including penicillins, cephalosporins, and carbapenems in poultry muscle using liquid chromatography coupled to tandem mass spectrometry. Anal Bioanal Chem 405:7859–7874
Camara M, Gallego-Pico A, Garcinuno RM, Fernandez-Hernando P, Durand-Alegria JS, Sanchez PJ (2013) An HPLC-DAD method for the simultaneous determination of nine beta-lactam antibiotics in ewe milk. Food Chem 141:829–834
Cerniglia CE, Kotarski S (1999) Evaluation of veterinary drug residues in food for their potential to affect human intestinal microflora. Regul Toxicol Pharm 29:238–261
Chen SP, Yu XD, Xu JJ, Chen HY (2011) Gold nanoparticles-coated magnetic microspheres as affinity matrix for detection of hemoglobin A1c in blood by microfluidic immunoassay. Biosens Bioelectron 26:4779–4784
Commission European (2010) Commission Regulation (EU) No 37/2010 of 22 December 2009 on pharmacologically active substances and their classification regarding maximum residue limits in foodstuffs of animal origin. Official Journal of the European Union 15:1–72
Fan F, Shen H, Zhang G, Jiang X, Kang X (2014) Chemiluminescence immunoassay based on microfluidic chips for alpha-fetoprotein. Clin Chim Acta 431:113–117
Fu Z, Yan F, Liu H, Yang Z, Ju H (2008) Channel-resolved multianalyte immunosensing system for flow-through chemiluminescent detection of alpha-fetoprotein and carcinoembryonic antigen. Biosens Bioelectron 23:1063–1069
He S, Zhang Y, Wang P, Xu X, Zhu K, Pan W, Liu W, Cai K, Sun J, Zhang W, Jiang X (2015) Multiplexed microfluidic blotting of proteins and nucleic acids by parallel, serpentine microchannels. Lab Chip 15:105–112
Kloth K, Rye-Johnsen M, Didier A, Dietrich R, Martlbauer E, Niessner R, Seidel M (2009) A regenerable immunochip for the rapid determination of 13 different antibiotics in raw milk. Analyst 134:1433–1439
Lamar J, Petz M (2007) Development of a receptor-based microplate assay for the detection of beta-lactam antibiotics in different food matrices. Anal Chim Acta 586:296–303
Lee M, Lee H, Ryu P (2001) Public health risks: chemical and antibiotic residues—review. Asian Austral J Anim 14:402–413
Lee TM, Carles MC, Hsing IM (2003) Microfabricated PCR-electrochemical device for simultaneous DNA amplification and detection. Lab Chip 3:100–105
Li H, Zhao M, Liu W, Chu W, Guo Y (2016) Polydimethylsiloxane microfluidic chemiluminescence immunodevice with the signal amplification strategy for sensitive detection of human immunoglobin G. Talanta 147:430–436
Macarov CA, Tong L, Martinez-Huelamo M, Hermo MP, Chirila E, Wang Y, Barron D, Barbosa J (2012) Multi residue determination of the penicillins regulated by the European Union, in bovine, porcine and chicken muscle, by LC-MS/MS. Food Chem 135:2612–2621
Märtlbauer E, Usleber E, Schneider E, Dietrich R (1994) Immunochemical detection of antibiotics and sulfonamides. Analyst 119:2543–2548
Pan X, Jiang L, Liu K, Lin B, Qin J (2010) A microfluidic device integrated with multichamber polymerase chain reaction and multichannel separation for genetic analysis. Anal Chim Acta 674:110–115
Perez B, Prats C, Castells E, Arboix M (1997) Determination of cloxacillin in milk and blood of dairy cows by high-performance liquid chromatography. J Chromatogr B 698:155–160
Schneider MJ, Mastovska K, Solomon MB (2010) Distribution of penicillin G residues in culled dairy cow muscles: implications for residue monitoring. J Agr Food Chem 58:5408–5413
Shamsi MH, Choi K, Ng AH, Wheeler AR (2014) A digital microfluidic electrochemical immunoassay. Lab Chip 14:547–554
Shryock TR (1999) Relationship between usage of antibiotics in food-producing animals and the appearance of antibiotic resistant bacteria. Int J Antimicrob Ag 12:275–278
Strasser A, Dietrich R, Usleber E, Märtlbauer E (2003) Immunochemical rapid test for multiresidue analysis of antimicrobial drugs in milk using monoclonal antibodies and hapten-glucose oxidase conjugates. Anal Chim Acta 495:11–19
Sun J, Xianyu Y, Jiang X (2014) Point-of-care biochemical assays using gold nanoparticle-implemented microfluidics. Chem Soc Rev 43:6239–6253
Taranova NA, Byzova NA, Zaiko VV, Starovoitova TA, Vengerov YY, Zherdev AV, Dzantiev BB (2013) Integration of lateral flow and microarray technologies for multiplex immunoassay: application to the determination of drugs of abuse. Microchim Acta 180:1165–1172
Usleber E, Lorber M, Straka M, Terplan G, Martlbauer E (1994) Enzyme-Immunoassay for the detection of isoxazolyl penicillin antibiotics in milk. Analyst 119:2765–2768
Whitesides GM (2006) The origins and the future of microfluidics. Nature 442:368–373
Zhang L, Feng Q, Wang JL, Sun JS, Shi XH, Jiang XY (2015) Microfluidic synthesis of rigid nanovesicles for hydrophilic reagents delivery. Angew Chem Int Edit 54:3952–3956
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We thank Ms. Jing Wu, Mr. Ulas Acaröz, and Mr. Christoph Kunas for their assistance.
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The study was funded by Zhonghechuang Investment Co., Ltd, China, Wuhan PriCells Biomedical Technology Co., Ltd, China, and the CAS/SAFEA International Partnership Program for Creative Research Teams.
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Wenbo Yu was financially supported by Zhonghechuang Investment Co., Ltd, China and Wuhan by PriCells Biomedical Technology Co., Ltd, China. Yiping Chen and Xingyu Jiang received research grants by the CAS/SAFEA International Partnership Program for Creative Research Teams. All other authors declare that they have no conflict of interest.
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Yu, W., Chen, Y., Knauer, M. et al. Microfluidic Chip-Based Immunoassay for Reliable Detection of Cloxacillin in Poultry. Food Anal. Methods 9, 3163–3169 (2016). https://doi.org/10.1007/s12161-016-0508-4
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DOI: https://doi.org/10.1007/s12161-016-0508-4