Reagentless and reusable electrochemical affinity biosensors for near real-time and/or continuous operation. Advances and prospects

Nowadays, there is increasing demand to overcome the major constraints involved in cumbersome and protracted procedures used in conventional laboratory methods for molecular detection and develop methodologies suitable to near real-time, continuous, and direct monitoring of the target analyte in the analyzed sample at decentralized settings. In this context, biosensors using electrochemistry as transduction technique are particularly attractive to monitor binding-induced changes in the rigidity of a redox-tagged nucleic acid or peptide-switching probes (switch-based biosensors) or a capture antibody attached to an interrogating electrode. Sensors based on biomolecular switches, DNAs, aptamers, and peptides that reversibly shift between two or more conformations (or conformational ensembles) in response to the binding of a specific target ligand have been applied to the determination of a wide range of relevant analytes (nucleic acids, proteins, small molecules, and ions) and are rapid (responding in minutes), sensitive, reagentless, easily reusable, and less prone to fouling issues. They are capable to report in near real time the target binding event without any subsequent processing step, directly in challenging samples or continuously in flowing samples, and even in vivo using affordable and miniaturization-compatible instrumentation. Only a few redox-labeled capture antibody–based immunosensing strategies have been reported to date for the reagentless and continuous electrochemical determination of relevant analytes.