Development and Optimization of a mAb-Based Indirect Competitive ELISA for Enrofloxacin Residue in Duck

Abstract:

This paper presents the generation of monoclonal antibodies (mAbs) with high specificity against enrofloxacin (ENR) through cell fusion procedures, and the development of a mAb-based indirect competitive ELISA (icELISA) method to detect ENR residue using one of these Hybridomas (clone 4B5-D6). Under the optimal experimental conditions, this assay exhibited a working range of 0.004-38 ng/mL with IC50 and LOD values of 0.4 and 0.002 ng/mL, respectively. Except for a high cross-reactivity (105.2%) to Ciprofloxacin, negligible cross-reactivity to the other compounds was observed. After optimization, 10% of methanol was used in the assay buffer and this ELISA system can tolerate acetonitrile not higher than 10%. Recovery studies indicate that an excellent correlation between concentration spiked and concentration determined was found, and the results also suggest this assay has the potential to be incorporated into a quantitative monitoring program for the rapid screening of ENR residue in poultry tissues.

• Periodical: Applied Mechanics and Materials (Volumes 48-49)
• Pages: 423-427
• DOI: https://doi.org/10.4028/www.scientific.net/AMM.48-49.423
• Online since: February 2011
• Authors: Jin Qing Jiang, Hai Tang Zhang, Heng Zhang Zhao, Kun Zhao, Zhi Yong Xu, Xin Yao Chang, Zi Liang Wang
• Keywords: Detection, Duck, Enrofloxacin, Indirect Competitive ELISA, Monoclonal Antibody

References

[1] Zhao S, Jiang H, Li X, etc, in: Simultaneous determination of trace levels of 10 quinolones in swine, chicken, and shrimp muscle tissues using HPLC with programmable fluorescence detection [J]. J Agric Food Chem, 2007, 55(10): 3829-3834.

DOI: 10.1021/jf0635309

[2] Delepine B, Hurtaud-Pessel D and Sanders P, in: Simultaneous determination of six quinolones in pig muscle by liquid chromatography-atmospheric pressure chemical ionization mass spectrometry [J]. Analyst, 1998, 123(12): 2743-2747.

DOI: 10.1039/a804911d

[3] Dufresne G, Fouquet A, Forsyth D, etc, in: Multiresidue determination of quinolone and fluoroquinolone antibiotics in fish and shrimp by liquid chromatography/tandem mass spectrometry [J]. J AOAC Int, 2007, 90(2): 604-612.

DOI: 10.1093/jaoac/90.2.604

[4] Wang Z, Zhu Y, Ding S, etc, in: Development of a monoclonal antibody-based broad-specificity ELISA for fluoroquinolone antibiotics in foods and molecular modeling studies of cross-reactive compounds [J]. Anal Chem, 2007, 79(12): 4471-4483.

DOI: 10.1021/ac070064t

[5] Zhao Y, Zhang G, Liu Q, etc, in: Development of a Lateral Flow Colloidal Gold Immunoassay Strip for the Rapid Detection of Enrofloxacin Residues [J]. J Agric Food Chem, 2008, 56(24): 12318-12142.

DOI: 10.1021/jf802648z

[6] Watanabe H, Satake A, Kido Y, etc, in: Monoclonal-based enzyme-linked immunosorbent assay and immunochromatographic assay for enrofloxacin in biological matrices [J]. Analyst, 2002, 127(1): 98-103.

DOI: 10.1039/b109427k

[7] Batty J. D, Beatty B. G and Wlanos W. G, in: Measurement of monoclonal antibody affinity by non-competitive enzyme immunoassay [J]. J Immunol Methods, 1987, 100(1-2): 173-179.

DOI: 10.1016/0022-1759(87)90187-6

[8] James W. G, in: Monoclonal antibodies: Principles and Practice [M]. New York: Academic Press, (1983).