Impedance Biosensor Utilizing a Si Substrate Deposited by Wet Methods

Abstract

We report an impedance biosensor utilizing a Si electrode created by wet chemical deposition atop 6061 Al alloy. The sensor electrode is created by galvanic/electroless Si deposition from an electrolyte containing 10 mM HF and 20 mM Na2SiF6 in 80 wt% formic acid, followed by antibody immobilization. The impedance response of the sensor electrode to increasing concentrations of peanut protein Ara h 1, a common food allergen, can be fit to an equivalent circuit containing three RC loops. The circuit element most sensitive to antigen binding is the charge transfer resistance, yielding a detection limit of 4 ng/mL. Biosensors that utilize electrochemical impedance spectroscopy have been employed with a wide variety of immobilized biomolecules, including antibodies, receptor proteins, aptamers, and ssDNA. These biomolecules must be immobilized atop a conductive and biocompatible substrate, which is most commonly accomplished by amide bond formation to carboxylate-terminated Au-thiol self-assembled monolayers. However, Au-thiol self-assembly chemistry has been reported to have inadequate stability for many applications, with a shelf life limited to days to weeks. In addition, most sensors need to be calibrated, which for antibodybased biosensors requires antibody unfolding. For this reason, durable chemistry for antibody immobilization is also needed for biosensor regeneration during such a calibration procedure. In addition to Au, other biocompatible substrate materials that have been employed for impedance biosensors include C, Si, Pt, Ti, and ITO. Si is intriguing as a biosensor substrate, since it is directly below C in the periodic table, so Si-C bonds are of comparable strength to C-C and Si-Si bonds. Additional advantages of Si substrates for biosensors include easier incorporation into ULSI devices and easier surface preparation relative to C. Room temperature combined galvanic and electroless deposition of compact Si films was recently reported from concentrated formic acid. Here these Si films are used for immobilization of the mouse monoclonal antibody to peanut protein Ara h 1, and subsequent impedance detection of the protein antigen. Peanuts are considered one of the most dangerous food allergens, with severe anaphylactic reactions causing over 100 fatalities in the United States alone each year. Nine possible allergens within peanuts have been identified, Ara h 1 to Ara h 8, and peanut oleosin, with Ara h 1 the most widely studied.