The stabilization of Au NP-AChE nanocomposites by biosilica encapsulation for the development of a thiocholine biosensor.

We report on the construction of an amperometric biosensor based on the immobilization of the enzyme acetylcholinesterase (AChE) onto gold nanoparticles (Au NPs). The active enzyme is covalently bound directly onto the surface of the Au NPs via a thiol bond. This immobilization provides increased stability and high electron-transfer between the colloidal Au NPs, the catalyst and the transducer surface. To further increase the biosensor stability by protecting the enzyme from denaturation and protease attack, a layer of biosilica was grown around the Au NP enzyme nanocomposite. All steps, i.e., the conjugation of the enzyme to the gold nanoparticles and the encapsulation into biosilica, are monitored and confirmed by ATR-FT-IR spectroscopy. The stabilizing effect of the entrapment was evaluated amperometrically, while the operation of the biosensor was monitored over a period of 4 months. The initial sensitivity of the biosensor was calculated to be 27.58 nA mM(-1) with a linear response to the concentration of the substrate in the range from 0.04 to 0.4 mM. It is thus shown that the biosilica nanocomposites doped with Au NPs-AChE conjugates create a system that provides both signal mediation and significant enzyme stabilization over the existing AChE biosensor. The biosensor had retained all its activity at the end of the 4 months, compared with the normal AChE biosensor whose activity reached 50% after only 42 days of operation.