Abstract
In this study, the electropolymerization of 4-hydroxyphenylacetic acid (4-HPA) over graphite electrodes (GE) was optimized, aiming its application as a functionalized electrochemical platform for oligonucleotides immobilization. It was investigated for the number of potential cycles and the scan rate influence on the monomer electropolymerization by using cyclic voltammetry technique. It was observed that the polymeric film showed a redox response in the region of +0.53/+0.38 V and the increase in the number of cycles produces more electroactive platforms because of the better electrode coverage. On the other hand, the decrease of scan rate produces more electroactive platforms because of the occurrence of more organized coupling. Scanning electron microscopy (SEM) images showed that the number of potential cycles influences the coverage and morphology of the electrodeposited polymeric film. However, the images also showed that at different scan rates a more organized material was produced. The influence of these optimized polymerization parameters was evaluated both in the immobilization of specific oligonucleotides and in the detection of hybridization with complementary target. Poly(4-HPA)/GE platform has shown efficient and sensitive for oligonucleotides immobilization, as well as for a hybridization event with the complementary oligonucleotide in all investigated cases. The electrode was modified with 100 cycles at 75 mV/s presented the best responses in function of the amplitude at the monitored peak current values for the Methylene Blue and Ethidium Bromide intercalators. The construction of the genosensor to detect a specific oligonucleotide sequence for the Mycobacterium tuberculosis bacillus confirmed the results regarding the poly(4-HPA)/GE platform efficiency since it showed excellent sensitivity. The limit of detection and the limit of quantification was found to be 0.56 (±0.05) μM and 8.6 (±0.7) μM, respectively operating with very low solution volumes (15 µL of probe and 10 µL target). The biosensor development was possible with optimization of the probe adsorption parameters and target hybridization, which led to an improvement in the decrease of the Methylene Blue (MB) reduction signal from 14% to 34%. In addition, interference studies showed that the genosensor has satisfactory selectivity since the hybridization with a non-specific probe resulted in a signal decrease (46% lower) when compared to the specific target.
Highlights
Organic compounds have been widely used for the development of organic thin films since the discovery of the polyacetylene polymerization by Shirakawa et al in 1977 [1]
The continuity of these delocalized orbitals insures different electronic properties from those attributed to common polymers, such as low ionization energy, electrical conductivity, and high electronic affinity [6], and have been widely used as electrochemical platforms for biosensors development due to their charge transport properties and biocompatibility in a single structure [7]
Electrochemical experiments for polymeric film generation and characterization were performed in an electrochemical cell with 25.0 mL capacity, while all the biomolecules detection experiments were performed in an electrochemical cell with 1.0 mL capacity
Summary
Organic compounds have been widely used for the development of organic thin films since the discovery of the polyacetylene polymerization by Shirakawa et al in 1977 [1]. The continuity of these delocalized orbitals insures different electronic properties from those attributed to common polymers, such as low ionization energy, electrical conductivity, and high electronic affinity [6], and have been widely used as electrochemical platforms for biosensors development due to their charge transport properties and biocompatibility in a single structure [7] These polymers have shown an excellent platform for biomolecules immobilization since they are relatively inexpensive materials, the production techniques are simple, they can be deposited onto different electrode materials and the choice of different molecular structures provide the construction of films with completely different characteristics [8,9]. These polymers have preserved functional groups that are responsible for optimizing the interaction with the biomolecules (functionalized platforms)
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