Signal amplification for DNA detection based on the HRP-functionalized Fe3O4 nanoparticles

An electrochemical approach for the sensitive detection of sequence-specific DNA has been developed. Horseradish peroxidase (HRP) assembled on the Fe3O4 nanoparticles (NPs) were utilized as signal amplification sources. High-content HRP was adsorbed on the Fe3O4 NPs via layer-by-layer (LbL) technique to prepare HRP-functionalized Fe3O4 NPs. Signal probe and diluting probe were then immobilized on the HRP-functionalized Fe3O4 NPs through the bridge of Au NPs. Thereafter, the resulting DNA-Au-HRP-Fe3O4 (DAHF) bioconjugates were successfully anchored to the gold nanofilm (GNF) modified electrode surface for the construction of sandwich-type electrochemical DNA biosensor. The electrochemical behaviors of the prepared biosensor had been investigated by the cyclic voltammetry (CV), chronoamperometry (i-t), and electrochemical impedance spectroscopy (EIS). Under optimal conditions, the proposed strategy could detect the target DNA down to the level of 0.7 fmol with a dynamic range spanning 4 orders of magnitude and exhibited excellent discrimination to two-base mismatched DNA and non-complementary DNA sequences.