(Keynote) Labeless Photoelectrochemical and Impedimetric Aptasensors and Immunosensors Based on Biofunctionalized Electropolymerized Polymers
Authors: Cosnier Serge , Marks Robert , Holzinger Michael , Gorgy Karine , Gondran Chantal , Le Goff Alan , Palomar Quentin , Kazane Imen , Haddache Fatima 
Published: February 2020

For four decades, the development of biointerfaces has been the subject of increasing research efforts in the field of biosensors and energy conversion. Owing to their high conductivity, inertness and electroactive surface area, graphene and carbon nanotube are widely used for the design of biosensors and their functionalization was successfully performed by electrogenerated polymers exhibiting affinity or covalent binding interactions towards biomolecules.
Recent examples of electropolymerized films will be presented for the design of labeless immunosensors and aptasensors for bisphenol A, cocaine, dengue antibody or cholera toxin antibody [1].
We report thus the synthesis and electrochemical characterization of a novel electropolymerizable Ru(II) complex bearing an intercalator group as well as the electrochemical behavior and photoelectrochemical properties of the resulting polymer film [2]. The latter was applied to the label-free photoelectrochemical detection of cocaine. A photoelectrode was thus designed by electrodeposition of a pyrrole monomer modified with a polypyridyl Ru(II) complex bearing benzo[i]dipyrido-[3,2-a:2’.3’-c]phenazine (dppn) ligand. Owing to the intercalating properties of these immobilized complexes towards DNA double helix, cocaine aptamer was immobilized on the modified electrodes thanks to its stem-loop configuration in order to design a photoelectrochemical cocaine aptasensor. The detection of cocaine via the formation of the aptamer/cocaine complex was successfully observed via a decrease in the photocurrent intensity. The photoelectrochemical aptasensor exhibits a detection limit of 10 nmol L-1 and linear range of 1 10-8 to 5 10-4 mol L-1 [3].
We report here the original synthesis of an anti-bisphenol A (BPA) aptamer conjugated at its 5’-end with a pentahistidine peptide and its use for the design of an impedimetric BPA aptasensor. The introduction of a histidine tag is dedicated to the non-covalent binding of the aptamer onto electrodes previously modified by polypyrrole nitrilotriacetic films. The aptamer immobilization occurs via the histidine axial ligation onto Cu complex chelated by the polypyrrole film. This immobilization method, already used by our group to elaborate immunosensors or DNA sensors, offers a close proximity at the molecular level between the immobilized probe and the modified electrode surface. This approach thus confers a high sensitivity to the detection process, namely the aptamer/BPA folding. The resulting labeless impedimetric aptasensor displays a wide linear range from 10-11 to 10-6 mol L-1 with a sensitivity of 26 W cm2 per decade and an excellent specificity towards interfering agents such as 4,4'-dihydroxybiphenyl and bisphenol P.
With the aim to improve the sensitivity of impedimetric biosensors, we have investigated the influence of a multiwalled carbon nanotube (MWCNT) coating and its thickness on the impedimetric transduction. We present thus a reliable procedure based on dispersion and filtration of MWCNT forming homogeneous films with controlled thicknesses and morphologies, which can easily be transferred on electrode surfaces. Polypyrrole-NTA films were electrogenerated on MWCNT modified glassy carbon electrode to immobilize biotinylated cholera toxin via the coordination of the biotin group onto Cu2+ complex chelated on the polymerized NTA groups. Concerning the design of this immunosensor, different thicknesses of carbon nanotube films were tested. For this purpose, three different volumes of CNT suspension solutions were filtered on a cellulose filter and then transferred to the electrode surface for modification of the three different thickness films by electropolymerization and post-biofunctionalization. The resulting impedimetric immunosensor for cholera toxin antibody, shows excellent reproducibility, increased sensitivities, and an exceptional linear range for antibody detection between 0.1 pg.mL-1 to 1 mg.mL-1. Similar results were obtained for the detection of Dengue antibody, namely a detection limit of 0.1 pg.mL-1.

References

F. Cecchini, L. Fajs, S. Cosnier, R. S. Marks, Trends in analytical chemistry, in press

F. Haddache, A. Le Goff, B. Reuillard, K. Gorgy, C. Gondran, N. Spinelli, E. Defrancq, S. Cosnier, Chem. Eur. J. 2014, 20, 15555-15560.

F. Haddache, A. Le Goff, N. Spinelli, P. Gairola, K. Gorgy, C. Gondran, E. Defrancq, S. Cosnier, Anal. Chem., submitted.