Development of ELISA-Like Fluorescence Assay for Melamine Detection Based on Magnetic Dummy Molecularly Imprinted Polymers

We present a directly competitive fluorescence assay for highly sensitive detection of melamine in milk using magnetic dummy molecularly imprinted polymers (MDMIPs). The detection principle is based on competitive binding between the fluorescent label and melamine on the MDMIPs. The fluorescent label was obtained by combining fluorescein isothiocyanate (FITC) with melamine in ethanol and water. MDMIPs were prepared on the surface of Fe3O4@SiO2 nanoparticles using 2,4-diamino-6-methyl-1,3,5-triazine as dummy template. The MDMIPs were characterized and their adsorption capacity was evaluated based on their static adsorption and Scatchard analysis. Results suggest that MDMIPs were successfully coated on the Fe3O4@SiO2 surface and had a core-shell structure. Adsorption experiments suggested that the MDMIPs had higher specific recognition capacities for melamine and FITC-melamine (FITC-Mel) than did magnetic dummy molecularly non-imprinted polymers. Competitive binding between FITC-Mel and melamine was performed under the optimum conditions to determine melamine quantitatively. The linear range of this fluorescence assay was 0.1-20 mg/L for melamine detection. The detection limit was 0.05 mg/L in negative milk samples. The assay was also successfully employed to detect melamine in spiked milk samples, with satisfactory recoveries, i.e., between 70.2% and 92.7%.