Immobilisation of proteins at silicon surfaces using undecenylaldehyde: demonstration of the retention of protein functionality and detection strategies

摘要

We describe a simple method for the covalent immobilisation of proteins to hydrogen-terminatednsilicon surfaces and demonstrate various protein detection strategies. Using hydrosilation chemistry,n1-undecenylaldehyde is attached to the surface through stable Si–C bonds; the reaction occursnprimarily via the vinyl group and mainly aldehyde groups are presented at the top surface of thenmonolayer. Proteins are then captured by reaction with their surface lysines. The proteins are bound viana Schiff base, whose formation is reversible, but can be fixed by reduction with cyanoborohydride inna one-pot reaction. Using standard methods of patterning, we were able to specifically localise proteinsn(urease, amyloid beta (Ab1–42), GFP and TolAIII-GFP) with little non-specific adsorption at non-nreactive sites.We characterised the immobilised proteins by X-ray photoemission spectroscopy, atomicnforce microscopy and FTIR, and showed that they retain their functionality using potentiometry,nfluorescence and coupled antibody systems with chromogenic substrates. We also exploited thenconductivity of the silicon substrate to demonstrate electrochemical detection of surface-boundnproteins. These protocols will aid the development of protein biochips based on silicon, which gives risento the possibility of detecting protein–protein and protein–small molecule interactions electronically.nSuch chips would be expected to be of utility for comparative proteomics and in molecular medicine,ndrug discovery and diagnostics.