The Detection Of Cow Milk Adulteration In Goat Milk By Colloidal Gold Immunochromatographic Strip

With the improvement of people’s living situation, the consumption of milk and dairy products is getting higher and higher. Among milk products, the goat milk is easily digested and absorbed by human body, can relieve people from most of the allergies caused by milk proteins and enhances people’s resistance to disease. Thus, the goat milk is increasingly favored by more consumers. However, the yield of the goat milk is low, and its price is high. So, driven by the interests, cow milk and inferior milk have been added to goat milk to reduce costs and seek high profit in the market. In order to safeguard consumers’ interests and health, meet requirements of rapid on-site detection in the industrial production, with bovine IgG as the target analyte, gold immunochromatography assay for the rapid detection of cow milk adulteration in goat milk was carried out in this study. The main research contents and results are as follows:(1) Preparation, purification and identification of goat-anti-bovine IgG polyclonal antibody(pAb). With standard bovine IgG(purity > 90%) as the immunizing antigen, three healthy male goats were immunized. After the last immunization, the antiserum were collected, purified and identified. Results showed that the purity of the pAb was more than 90% after purification, the titer identified by indirect ELISA method was 1:1.28×105,the sensitivity could reach to 156.25 ng/mL and the specificity was good. These results demonstrated that the pAb could provide a necessary foundation for the development of the colloidal gold immunochromatography test.(2) Preparation, purification and idntification of mouse-anti-bovine IgG monoclonal antibody(McAb). After fusion of spleen cells and myeloma cells SP2/0, five positive clones were successfully screened out. Then the best clone of 3A4 was used to inoculate into abdominal cavity of the Balb/c mice for large-scale preparation of McAb.The identification results by ELISA methods showed that the titer of 3A4 McAb was 1:5.12× 105, the sensitivity was 78.125 ng/mL and the affinity constant was 5.73 × 108 L/mol.The cross reaction test showed that 3A4 McAb possessed a strong specificity to the bovine IgG.(3) Development and performance evaluation of the gold immunochromatography assay(GICA). A sandwich analysis model of pAb-target-McAb was used for the construction of GICA in this study. The optimum condition of immunoreagent and assay were achieved. The visible detection limit of the GICA in real samples was 0.4 mg/L for bovine IgG adulterated in goat milk. The cross reaction tests results of the developed GICA for α-casein, β-casein,α-lactoglobulin, β-lactoglobulin and goat IgG were negative. The coincidence rate of the sample addition experiment was 100%. These results indicate that the test strips can be applied to the rapid detection of cow milk adulteration in goat milk in real seamples.