The Study On The Immunoassay Of The Residues Of Olaquindox And Its Main Metabolite

The safety of animal derived food has become a hot problem in the food safety industry, and all walks of life in the society are paying close attention to it. Residue of Olaquindox (OLA) and its main metabolite methyl-3-quinoxaline-2-carboxylic acid (MQCA) are harmful to human health. The paper aims to research on determination of OLA and MQCA by immunoassay.The derivatized OLA was coupled to bovine serum albumin (BSA), Ovalbumin (OVA) to form immunogen and coating antigen. After immunization, a specific antibody against OLA was got and an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) was established. The 50% inhibitory concentration (IC50) was 151.89ng/mL and the limit of detection (LOD) was 4.33ng/mL. A novel synthesis method of methyl-3-quinoxaline-2-carboxylic acid (MQCA) and its hapten preparation were described. MQCA was synthesized involving two steps with a high purity. With the aim to improve the sensitivity of detection, four different haptens were synthesized to optimize the most suitable immunogen and coating antigen. After comparing the sensitivity of immunoassay, a specific antibody immunized with hapten (4-atom spacer arm)-BSA and a desirable coating antigen with a heterologous spacer arm group were obtained. Using the designated antibody and coating antigen, a degree of class selectivity as a first-pass screening assay was developed and the sensitivity of ic-ELISA showed that the IC50 was 3.84ng/mL and the LOD was 0.25 ng/mL.An ultra-sensitive, rapid, high-throughout fluorescence-quenching assay for small molecuLar veterinary drug residue detection was developed by using nanoparticle signal amplification. This system uses a gold nanoparticle probe conjugated with goat anti-rabbit IgG and DNA as a substitute of chemiluminescent probe in sequential injection analysis (SIA) system. After an indirect-competitive immunoassay in SIA system, the DNA functionalized gold nanoparticle probe attached to superparamagnetic nanoparticles immobilized with antibody-antigen complex. The DNA on the gold nanoparticle was dehybridized and the single strand DNA (ssDNA) was then hybridized off-line with another smaller-sized complementary oligonucleotide functionalized gold-nanoparticles to amplify the signal, followed with a sensitive fluorescence quenching detection. This assay showed a lower limit of detection (7.52 pg/mL) as well as a desirable range from 8.5 to 60 pg/mL that couLd be extended to a potential broad biomedical analysis.