Characterization of biotin-avidin recognition system constructed on the solid substrate

Abstract

The biotin-avidin complex, as a model recognition system, has been constructed through N-hydroxysuccinimide (NHS) reaction on a variety of substrates such as a smooth Au film, electrochemically roughened Au electrode and chemically modified mica. Stepwise self-assembled monolayers (SAMs) of biotin-avidin system were characterized by surface-enhanced resonance Raman scattering (SERRS) spectroscopy, atomic force microscopy (AFM) and surface plasmon resonance (SPR). A strong SERRS signal of rhodamine tags labeled in avidin from the SAMs on a roughened gold electrode indicated the successful complex formation of stepwise biotin-avidin recognition system. AFM images showed the circular shaped avidin aggregates (hexamer) with ca. 60 Å thick on the substrate, corresponding to one layer of avidin. The surface coverage and concentration of avidin molecules were estimated to be 90% and 7.5×10−12 mol/cm2, respectively. SPR technique allowed one to monitor the surface reaction of the specific recognition with high sensitivity and precision.