Attachment of ferrocene-capped gold nanoparticle-streptavidin conjugates onto electrode surfaces covered with biotinylated biomolecules for enhanced voltammetric analysis

Ferrocene (Fc)-capped gold nanoparticle/streptavidin conjugates were attached onto electrodes covered with either biotinylated oligodeoxynucleotides (ODNs) duplexes or N-biotinoyl-N'-(6-maleimidedohexanoyl)hydrazide that had been preadsorbed onto the reduced form of glutathione (GSH) immobilized onto a dithiobissuccinmidyl propionate self-assembled monolayer (SAM). Effects of the DNA probe and target strand lengths on the voltammetric behavior of the Fc groups residing at the nanoparticle conjugates were studied. The voltammetric behavior was also compared to that arising from the Fc groups present at the end of the ODN duplex in a sandwich complex (Wang et al. Anal. Chem. 2003, 75, 3941). It was found that shorter duplexes would lead to a more reversible voltammetric wave. However, the formation of duplexes with a greater number of base pairs will result in a lower detection level. For the analysis of a biotinylated 30mer target at a mixed SAM of a 30mer probe and hexanethiol, a concentration level as low as 0.25 pM could be detected (which is even lower than our previously reported detection level of 2 pM for an ODN sandwich complex). The detection level for GSH (1 nM) was found to be comparable to those of many other analytical techniques. The capability of our method for trace analysis of biotinylated biomolecules can be attributed to the enhanced voltammetric detection by the large number of Fc moieties present at each conjugate. The method described herein is simple, reproducible, versatile, and selective, and does not require sophisticated analytical instrumentation.