Characterization and optimization of a real-time, parallel, label-free, polypyrrole-based DNA sensor by surface plasmon resonance imaging

We describe in this paper a methodology to quantify multispot parallel DNA hybridizations and denaturations on gold surfaces by using, on one hand, a polypyrrole-based surface functionalization based on an electrospotting pro cess and, on the other hand, surface plasmon resonance imaging allowing rea l-time measurements on several DNA spots at a time. Two characterization st eps were performed in order to optimize the immobilization of oligonucleoti de probes and, thus, to increase the signal-to-noise ratio of monitored hyb ridization signals: the first step consisted of characterizing the signal d ependence upon the density of immobilized 15-mer probes, and, the second st ep, in analyzing the hybridization response versus spot thickness. We furth er demonstrated that a surface density of polypyrrole/DNA probes of similar to 130 fmol/ mm(2) (590 pg/mm(2)) optimizes the hybridization signal that can be detected directly. Optimal thickness of the spot was found to be clo se to 11 nm. Specificity and regeneration steps on each spot have also been demonstrated successfully, showing this method to be very competitive and convenient in use.