Rethinking the use of redox probes for the detection of electroactive proteins with electrochemical sensors modified with molecularly imprinted polymers

This study aims to demonstrate that redox couples, regardless of their electrical charges, are unnecessary for detecting and quantifying electroactive proteins using an electrochemical sensor functionalized with a molecularly imprinted polymer. Our approach involved designing a polydopamine imprinted biosensor for detecting bovine serum albumin as the model protein. Electrochemical measurements were conducted in a phosphate-buffered solution (PBS) and solutions containing the negatively charged hexacyanoferrate, the neutral ferrocene, or the positively charged hexaammineruthenium (III) probes. The dissociation constants Kd (in mg.mL−1), estimated from an extended Langmuir/one-site model, were of order of (1.0 ± 0.5)×10−8, (4.4 ± 2.1)×10−2, (7.6 ± 5.1)×10−4 and in the presence of [Fe(CN)6]−3/−4, Fe(C5H5)2, [RuN6H18]+3 respectively, and (8.7 ± 5.9)×10−11 in PBS. The non-use of probes, therefore, enhances the interaction between the analytes and the imprints. To understand the origin of this finding, we investigated ultraviolet and Fourier-transform infrared spectroscopies. Results indicated that redox probes could alter the proteins' intrinsic properties and adsorb to the polydopamine polymeric matrix, thus reducing the specific interactions between the protein and the imprints. To confirm the feasibility of electrochemical quantification of electroactive proteins in PBS, we designed three polydopamine-imprinted biosensors for detecting human serum albumin, prostate-specific antigen, and immunoglobulin G. Results validated the potential for quantifying electroactive proteins in PBS without adding any probe. This pioneering study was carried out with dopamine, which is taken here as a typical example of a functional monomer. It paves the way towards the detection of electroactive proteins without adding any redox couple of any nature.