Qualitative and quantitative detection of DNA amplified with HRP-modified SiO₂ nanoparticles using scanning electrochemical microscopy

Qualitative and quantitative detection of DNA was achieved by a "sandwich" DNA sensor through SG/TC (substrate generation and tip collection) mode of scanning electrochemical microscopy (SECM). The "sandwich" DNA structure was formed by the hybridization of thiol-tethered oligodeoxynucleotide probes (capture probe), assembled on the gold substrate surface, with target DNA and biotinylated indicator probe. HRP (horseradish peroxidase)-wrapped SiO₂ nanoparticles were linked to the sandwich structure through biotin-streptavidin interaction. Hydroquinone (H₂Q) was oxidized to benzoquinone (BQ) at the modified substrate surface where sequence-specific hybridization had occurred through the HRP-catalyzed reaction in the presence of H₂O₂. The detection was based on the reduction of BQ generated on the modified substrate by SECM tip. For SECM imaging experiment, we structured the microsensor platform through localized desorption of 1-dodecanethiol monolayer. Approach curves were employed for quantitative detection of DNA concentration. The detection limit of complementary DNA was as low as 0.8pM. This technique is promising for the application on electrochemical DNA chip.