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Xu Y.-X.,China Jiliang University | Gong Y.-F.,China Jiliang University | Chen Z.-L.,Hangzhou DNA Science and Technology Co. | Xi X.,China Jiliang University | And 4 more authors.
Modern Food Science and Technology | Year: 2014

To develop immunoassay for gentamicin, the artificial antigen and polyclonal antibody were prepared in this study. Identified through ultraviolet spectrum (UV) and SDS-PAG, the immunogen (GM-BSA) and coating antigen (gm-ova) of gentamicin were synthesized by the method of carbodiimide successfully. GM-BSA was used to immune New Zealand white rabbits, and the antiserum was obtained. In this study, we utilize a notive method to immune animal and obtained serum volume much more than conventional method's. One indirect competitive ELISA method has been developed to detect the titer and sensitivity of the antiserum. The results show that the titer of antiserum was continuously improving and the IC50 was nearly 3 ng/mL. On this basis, With an concentration range from 0.3 to 243 ng/mL, the IC50 is 2.954 ng/mL, and the IC10 is 0.056 ng/mL. The results of this study provide a basis for the breakthrough of serum volume, and also is a good start for the development of gentamicin ELISA kit. Source

Gong Y.,China Jiliang University | Zhang M.,China Jiliang University | Wang M.,Hangzhou DNA Science and Technology Co. | Chen Z.,Hangzhou DNA Science and Technology Co. | Xi X.,China Jiliang University
Arabian Journal for Science and Engineering | Year: 2014

A competitive indirect enzyme-linked immunosorbent assay (ELISA) and a colloidal gold-based immunochromatographic assay (CGIA) using monoclonal antibodies (mAbs) were developed for the detection of melamine. Under optimum conditions, IC50 value was 8 and 8.3 ng/mL for two mAbs (2H9 and 1B12), respectively. The ELISA kit based 1B12 showed detection limit of 1.908 ng/mL, 2.125 and 4.154 ng/mg for milk, milk powder and animal feed samples, respectively; while that of 2H9 exhibited detection limit of 5.210 ng/mL, 5.748 and 5.460 ng/mg for the same products, respectively. No cross-reactivity was found from the other tested compounds. Milk, milk powder and animal feed samples spiked with concentration range from 0 to 500 ng/mL were analyzed by the established ELISA. Acceptable recovery rates of 84.74-104.94, 84.52-104.05 and 83.3-94.67 % were obtained for milk, milk powder and animal feed samples, respectively. The optimized CGIA-based 2H9 was utilized in the detection of melamine in milk, milk powder and animal feeds, with detection limit of 100 ng/mL for milk, 100 ng/mg for milk powder and 200 ng/mg for feeds, respectively. The ELISA and CGIA procedures established in this study allow on-site screening of melamine residues in food samples. © 2014 King Fahd University of Petroleum and Minerals. Source

Gong Y.,China Jiliang University | Chen Z.,Hangzhou DNA Science and Technology Co. | Xi X.,China Jiliang University | Li M.,China Jiliang University | And 3 more authors.
Shengwu Gongcheng Xuebao/Chinese Journal of Biotechnology | Year: 2012

To develop a specific, rapid, and convenient immunochromatography assay (ICA) to detect melamine residues in dairy products and feed sample. Colloidal gold particles labeled with purified monoclonal antibody against anti-melamine were used as the detector reagent. The MEL-OVA (the conjugate of melamine and ovalbumin) and goat anti-mouse melamine IgG were blotted on the test and control regions of nitrocellulose membrane. The strip was then assembled with sample pad, absorbing pad, and dorsal shield. The limit of detection (LOD) is 50 μg/L. The test trip was applied to detect melamine in milk, milk powder, and animal feeds, with detection limits of 100 μg/L for milk, 100 ng/g for milk powder, 200 ng/g for feeds. Compared to LC-MS/MS, the ICA could be used to screen a large number of dairy products and feed samples for melamine residue. Source

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