Estudo para o desenvolvimento de biossensor impedimétrico não marcado para detecção de veneno ofídico

Abstract

Snakebites represent a serious public health problem due to the frequency and severity of the accidents all over the world, and serotheraphy is the only approved treatment for envenomed patients. However, the right serum management to the snakebite patient is still performed using inconsistent techniques. In this sense, this research purposed the study of biosensors development for analytical and specific detection of Bothrops snakes venom, genus responsible for the major occurrence of snakebites in Brazil. Thus, the employment of different transducer substrates was evaluated for the immunodiagnosis development, producing Polyaniline (PANI) and titanium dioxide, and commercially obtaining substrates of tin-doped indium oxide, Crofer 22 APU steel and gold, which were characterized by electrochemical, spectroscopic and microscopic techniques. To functionalize these substrates, Immunoaffinity Chromatography was used to purify antibodies from plasma of horse immunized against a pool of venoms from B. alternatus, B. jararaca, B. jararacussu, B. moojeni e B. neuwiedi, whose purity was revealed by Polyacrylamide Gel Electrophoresis. The obtained antibodies were employed as element of biological recognition onto the transducers surface and the produced biosensors were evaluated for their sensibility and specificity for detection of bothropic venom, using Electrochemical Impedance Spectroscopy, in a frequency range of 104 a 10-2 Hz, with a potential amplitude of 10 mV above the open circuit potential. The results indicated the efficacy of PANI and TiO2 production, the success of the transducers functionalizing and their capability to detect the analyte of interest in an average time of 36.58 minutes. PANI showed the greatest analytical sensibility (with a minimum limit detection lower than 0,1 g.mL-1) and a high specificity, not generating a significant impedimetric response when it was exposed to venom from snakes ofCrotalus and Micrurus genera.