Desenvolvimento de biossensores eletroquímicos, utilizando matriz polimérica modificada com biomoléculas, para o diagnóstico de câncer de próstata e hepatite C

Abstract

The development of biologic sensors has become a wide activity in science seeking mainly health and environmental issues. The biosensor development in laboratory has been investigated by many researchers seeking sensitivity, specific, low cost and portable devices. The electrochemical techniques are used in the largest number of works due to the easiness in the process and in the instrumentation, allowing for the rapid and reliable results. Modified electrodes with conducting polymers have been used because they improve the analytical response in addition to aid in the biomolecule immobilization. This work aimed the use of graphite electrodes and graphite electrodes modified with poly(3-aminophenol) for the construction of DNAbased biosensors for the detection of hepatitis C virus and prostate cancer. The polymer presents adequate conditions (ease in formation, thermal and electrochemical stability besides the presence of functionalizations in its structure) for the sensor construction. The denatured PCR products were successfully immobilized over the polymeric matrix. The hybridization indirect detections were performed using ethidium bromide (EB) oxidation signal. The detection system for prostate cancer and hepatitis C was optimized with 15 minutes of interaction between target and probe, concentration of EB at 0.54 mmol.dm-3, detection limit of 16 and 19 ng and quantification limit of 54 and 64 ng for prostate cancer and hepatitis C, respectively and lifetime for the sensors of 60 days. Electrochemical impedance spectra were obtained proving the probe and target immobilization efficiency due to electrostatic differences between the anionic solution and the immobilized material. Increase in resistance and capacitance values of the systems containing the probe for the systems containing the targets shown a response difference, qualitatively. A mathematic model based on factorial design planning between EB and Calf Thymus DNA was performed through fluorescence spectroscopy, UV-visible spectroscopy and differential pulse voltammetry. A proportion of 6:1 for DNA/EB was found the best condition and, in the working range of this study, the influence of sodium chloride was investigated, with the better results for a concentration of 20 mmol.dm-3.