Carboxylic acid-tethered polyaniline as a generic immobilization matrix for electrochemical bioassays

Polyaniline (PANI) was functionalized by thiol-ene click chemistry to obtain carboxylic acid-tethered polyaniline (PCOOH). The versatility of PCOOH as an immobilization matrix was demonstrated by constructing four different biosensors for detection of metabolites and cancer biomarker. Immobilization efficiency of PCOOH was investigated by surface plasmon resonance and fluorescence microscopic analysis which revealed dense immobilization of biomolecules on PCOOH as compared to conventional PANI. A sandwich electrochemical biosensor was constructed using PCOOH for detection of liver cancer biomarker, α-fetoprotein (AFP). The sensor displayed sensitivity of 15.24 µA (ng mL^-1)^-1 cm^-2, with good specificity, reproducibility (RSD 3.4%), wide linear range (0.25-40 ng mL^-1) at - 0.1 V (vs. Ag/AgCl), and a low detection limit of 2 pg mL^-1. The sensor was validated by estimating AFP in human blood serum samples where the AFP concentrations obtained are consistent with the values estimated using ELISA. Furthermore, utilization of PCOOH for construction of enzymatic biosensor was demonstrated by covalent immobilization of glucose oxidase, uricase, and horseradish peroxidase (HRP) for detection of glucose, uric acid, and H₂O₂, respectively. The biosensors displayed reasonable sensitivity (50, 148, 127 µA mM^-1 cm^-2), and linear ranges (0.1-5, 0.1-6, 0.1-7 mM) with a detection limit of 10, 1, and 8 µM for glucose, uric acid, and H₂O₂, respectively. The present study demonstrates the capability of PCOOH to support and enable oxidation of H₂O₂ generated by oxidase enzymes as well as HRP enzyme catalyzed reduction of H₂O₂. Thus, PCOOH offers a great promise as an immobilization matrix for development of high-performance biosensors to quantify a variety of other disease biomarkers. Carboxylic acid-tethered polyaniline synthesized by thiol-ene click chemistry was used as matrix to construct four different electrochemical biosensors for detection of cancer biomarker α-fetoprotein, glucose, uric acid, and H₂O₂.