Development of Immuno-Based Methods for Detection of Melamine

Abstract

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.