Zhifeng Qin

Development and evaluation of an immunochromatographic strip for the detection of serum antibodies against bluetongue virus

In this study, an immunochromatographic strip (ICS) was developed for the detection of bluetongue virus (BTV) serum antibodies. Colloidal gold particles labeled with streptococcal protein G (SPG), which can bind to the F(C) fragment of mammalian immunoglobulins, were used as the detector reagent. A recombinant VP7 BTV protein and a purified rabbit anti-SPG antibody were immobilized on test and control regions of a nitrocellulose membrane, respectively. In order to evaluate the ICS, 37 sera from animals exposed to different BTV serotypes were used as positive controls. In addition, 50 positive sera against viruses other than BTV, and eight sera taken from naive healthy sheep were used to determine the specificity of the ICS. Three hundred and three field sera taken from sheep and cattle were used after the above sera had been used for validation. Compared with the competitive ELISA (c-ELISA), the specificity and sensitivity of the ICS was 97.6% and 100%, respectively. There was excellent agreement between the results obtained by c-ELISA and the ICS (kappa=0.930). As it is rapid and easy to use, the test is suitable for the serological surveillance of BTV infection in the field.

Simultaneous Screening for Five Veterinary Drug Residues by a Developed Multiplexed Luminex Assay

Abstract The development of sensitive and rapid methods for simultaneous determination of multiple residual veterinary drugs in foods is increasingly important. In this study we reported a new method to simultaneously detect five veterinary drug residues, including ractopamine, clenbuterol, furazolidone, chlortetracycline and qlaquindox. The experimental results indicated that the developed multiplex assay enable to detect all the five drugs at the concentration of 0.1 ng/mL, with desirable specificity, suggesting that it could be applicable for rapid screening of five residual veterinary drugs in a faster turnaround time and higher throughput.

Subtyping animal influenza virus with general multiplex RT-PCR and Liquichip high throughput (GMPLex)

This study developed a multiplex RT-PCR integrated with luminex technology to rapidly subtype simultaneously multiple influenza viruses. Primers and probes were designed to amplify NS and M genes of influenza A viruses HA gene of H1, H3, H5, H7, H9 subtypes, and NA gene of the N1 and N2 subtypes. Universal super primers were introduced to establish a multiplex RT-PCR (GM RT-PCR). It included three stages of RT-PCR amplification, and then the RT-PCR products were further tested by LiquiChip probe, combined to give an influenza virus (IV) rapid high throughput subtyping test, designated as GMPLex. The IV GMPLex rapid high throughput subtyping test presents the following features: high throughput, able to determine the subtypes of 9 target genes in H1, H3, H5, H7, H9, N1, and N2 subtypes of the influenza A virus at one time; rapid, completing the influenza subtyping within 6 hours; high specificity, ensured the specificity of the different subtypes by using two nested degenerate primers and one probe, no cross reaction occurring between the subtypes, no non-specific reactions with other pathogens and high sensitivity. When used separately to detect the product of single GM RT-PCR for single H5 or N1 gene, the GMPLex test showed a sensitivity of 10−5(= 280ELD50) forboth tests and the Luminex qualitative ratio results were 3.08 and 3.12, respectively. When used to detect the product of GM RT-PCR for H5N1 strain at the same time, both showed a sensitivity of 10−4(=2800 ELD50). The GMPLex rapid high throughput subtyping test can satisfy the needs of influenza rapid testing.