Facile electrochemical biosensor based on a new bifunctional probe for label-free detection of CGG trinucleotide repeat.

We have developed a simple and label-free electrochemical assay to detect CGG trinucleotide repeat. For this purpose, a new bifunctional probe (FecNCD2) was developed, in which a recognition part (naphthyridine carbamate dimmer, NCD) was connected with an electro-active part (ferrocenyl group) using a chain of -CO-NH-CH2-CH2-. The results of circular dichroismic measurements indicated that FecNCD2 exhibited a superior performance for selective binding to CGG trinucleotide repeats compared to a previous bifunctional electrochemical probe connected with shorter linker -CH2- (FecNCD1). Then, the electrochemical properties of FecNCD2 were evaluated and were found to show a good redox response due to the ferrocene moiety. Owing to the high performances of FecNCD2, the label-free electrochemical biosensor for CGG repeats was constructed by immobilizing them onto gold disk electrode and by using FecNCD2 as an electrochemical probe in solution. Further CGG repeats in solution were confirmed to be detectable using the CGG modified biosensor in competitive experiments, i.e., by treating it in test solutions containing FecNCD2 and d(CGG)10 or others. No interference of ct-DNA on the CGG detection was also confirmed with this approach. The strategy should have significant potential for the development of versatile and low-cost biosensor for early diagnosis and treatment of neurodegenerative diseases associated with trinucleotide repeats.

Selective recognition of G-G mismatch using the double functional probe with electrochemical active ferrocenyl.

A new electrochemical assaying approach with label-free to identify GG mismatch in DNA duplexes was developed by applying a new double functional probe (FecNC), ferrocenyl modified naphthyridine derivative, which contain recognition domain (naphthyridine derivative) and electroactive center (ferrocenyl group). The double functional probe exhibited not only the excellent electrochemical response devoted from ferrocenyl but also high selective electrochemical signal "off" for G-G mismatch duplex. The interaction was also verified by melting temperature and circular dichroismic spectra (CD). The electrochemical investigation showed that the redox of FecNC was a diffusion-controlled process and the diffusion coefficient of bound-FecNC was much smaller than free FecNC. The correlation between the current of FecNC and concentration of GG mismatch duplex indicated that the bind saturation point was about at a 1:1 mole ratio. The novel double functional electrochemical probe might provide a versatile and low-cost way to detect single nucleotide polymorphisms, which could be found extensive applications in the diagnosis of the genetic diseases.