Silicon nanoparticles coated with an epitope-imprinted polymer for fluorometric determination of cytochrome c

The authors describe a composite consisting of silicon nanoparticles that were first coated with SiO2 and then with a molecularly imprinted polymer (SiNP@SiO2@MIP). The MIP was generated by dual epitope imprinting such that it can recognize cytochrome c (Cyt c). The MIP on the NPs was prepared from the functional monomer zinc(II) acrylate (ZnA), the crosslinker ethylene glycol dimethacrylate and the initiator 2,2'-azoisobutyronitrile. Dual epitope templates for Cyt c included (a) a C-terminal nonapeptide (AYLKKATNE), and (b) an N-terminal nonapeptide (GDVEKGKKI). The chelation between Zn(II) of ZnA and the amino groups or hydroxy groups of the template nonapeptides warrants good recognition and capture of Cyt c. The fluorescence originating from SiNPs has excitation/emission peaks at 360/480 nm and is quenched by Cyt c in the 0.50-40.0 mu M concentration range. The correlation coefficient for the calibration plot of the imprinted NPs is 0.9937. The detection limit is 0.32 +/- 0.01 mu M, the precisions of six replicate detections at levels of 0.5, 20 and 40 mu MCyt c are 3.2, 2.7 and 2.8%, respectively, and the imprinting factor is 2.43. Compared to single epitope template imprinting, dual epitope imprinting results in improved selectivity. The imprinted nanoparticles can discriminate Cyt c even if one amino acid is mismatched. The method was applied to the determination of Cyt c in spiked diluted human serum and gave recoveries between 94.0 and 107.5%.