Controllable Nano-gap Biosensors For The Sensitive Electrochemical Detection Of Biological Macromolecules

Objective:Controllable nano-gap biosensors which are constructed based on interdigitated array microelectrode (IDAM) for streptavidin (SAv) and human chorionic gonadotropin (HCG) no immunolabeling sensitive detection research by using Electrochemical Impedance Spectroscopy.Methods:About 16 nm gold nanoparticles (AuNPs) in size are successfully prepared, through the simple hydrothermal synthesis method. The synthesized gold nanoparticles solution was characterized by UV-visible spectrophotometer (UV-vis) and environmental scanning electron microscope (SEM). Using the maximum surface plasmon resonance (SPR) absorption peaks of UV-visible spectrum and FIG environmental scanning electron microscope to calculate the AuNPs particle size and observe morphology.By chemical deposition method, we constructed novel nano-gap electrodes, so that the electrode gap shortened from micron to nanometer. At first, make the interdigitated array microelectrode silanized. Then, the gold nanoparticles were chemically bonded on the 2.5 μm electrode gap of the interdigitated array microelectrode. The electrode surface and the electrode gap before and after the deposition of gold nanoparticles was analyzed with the scanning electron microscope and the overall morphology of the electrode were characterized by metallurgical microscope.The nanogap biosensor with the optimum gap was manufactured through the in situ growth of gold nanoparticles which can adjust the particle size of the gold particles in the interdigitated array microelectrode gap for regulated the size of the gap between the gold nanoparticles.The electrode was characterized by the electrochemical methods including electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). And these two methods were used to study the detection effect of the electrode with different gap sizes. Finally, Target biomolecules (SAv, HCG) were detected by further application of EIS and CV. With the concentration increase of cysteamine-biotin-streptavidin and cysteamine-antibodies-bovine serum albumin (BSA)-antigen self-assembled on the electrode surface, the electrochemical properties of the electrode surface were changed.Results:Spherical gold nanoparticles of uniform size (about 16 nm) were successfully prepared. The uniform sized gold particles which are distributed orderly in the ID AM gap can be seen from scanning electron microscope.The nano-gap biosensor with suitable gap was constructed successfully by combining the approach of the in situ growth of gold nanoparticles.The electrode electrochemical characterization showed the biomolecules assembly process. When the gold nanoparticles were deposited on the ID AM, EIS is reduced significantly, and the cyclic voltammetry current increased obviously. And with cysteamine-biotin-streptavidin and cysteamine-antibodies-bovine serum albumin (BSA)-antigen self-assembled on the electrode surface gradually, EIS is increased gradually, and the cyclic voltammetry current decreased step by step. Compared with the detection effect of the other electrodes with different, the nano-gap electrode of 25 nm gap exhibited an excellent detection results.This novel sensor was successfully applied to determine biomolecules with high sensitivity (the detection limit is 1 pM), good selectivity and stability. EIS increased with the increasing of the target molecules concentration gradually (1～1×105 pM of streptavidin; 0.1～100 mlU/ml of antigen), while the corresponding cyclic voltammetry current gradually decreased. The concentration (C) and the absolute value of the electron transfer impedance (△Ret) shows good linear relationship. Linear regression equation was obtained.Conclusion:The streptavidin and human chorionic gonadotropin were detected rapidly and sensitivly by this nano-gap electrochemical biosensor combined. Electrochemical impedance spectroscopy and nano-gap electrochemical biosensor were combined for the first time in this study.