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Here we have demonstrated a general, sensitive, and selective approach for the detection of macromolecules that bind to specific small molecule recognition elements. Our electrochemical approach utilizes a redox-tagged DNA signaling scaffold that is conjugated to a small molecule recognition element and is covalently attached to an interrogating electrode. The binding of a protein to the small molecule recognition element alters the dynamics of the scaffold, increasing or decreasing the efficiency with which the redox tag collides with the electrode and thus altering the observed faradaic current. We optimized the scaffold using a biotin recognition element and streptavidin as a target to determine the variables that define sensor performance before then applying the approach to detection of anti-digoxigenin antibodies using the steroid as the recognition element. We generated streptavidin sensors exhibiting both signal-on (target binding increases the faradaic current) and signal-off behavior, of which only the signal-off approach was generalizable to the detection of antibodies. Sensors for both targets are sensitive (detection limits in the low nanomolar range), rapid (minutes), reusable, and selective enough to function directly in complex matrices including blood serum, soil, and foodstuffs.

An Electrochemical Sensor for the Detection of Protein−Small Molecule Interactions Directly in Serum and Other Complex Matrices