Desenvolvimento e caracterização de imunossensor eletroquímico para detecção de câncer de tireoide diferenciado

Abstract

Electrochemical immunosensors applied in the biomedical field are devices built based on high sensitivity and precision methods, fundamental requirements for monitoring biomarkers in patients undergoing treatment for differentiated thyroid cancer. Polymeric nanocomposites are promising alternatives for biosensor production due to their unique properties. In this work, an electrochemical immunosensor was developed and characterized the use of polymeric nanocomposites as a platform for the immobilization of biotinylated anti-thyroglobulin (anti-Tg) antibodies for the detection of thyroglobulin (Tg). First, three polymeric matrices were used to produce nanocomposites, such as cellulose acetate, EuDragit-100® and poly(vinylidene) fluoride (PVDF) containing TiO2 nanoparticles. The PVDF-based nanocomposite presented better electrochemical profile, so three more nanocomposites were prepared using PVDF containing TiO2 nanotubes, graphene oxide and gold nanoparticles, respectively. A preliminary electrochemical study was performed related to the immobilization of DNA and serum albumin (BSA) structures on TiO2/ITO/glass semiconductor electrodes in order to evaluate the immobilization of these biological structures on TiO2, using electrochemical and spectroscopic techniques photoemission (XPS). For the biotinylated anti-Tg immobilization platform, streptavidin-coated iron oxide microparticles (MPs) were incorporated into the polymer matrix for the formation of the streptavidin-biotin system. Nanocomposite films were obtained by spin coating, electrospinning, and casting, employing four electrodes: ITO electrode, gold electrode, glassy carbon electrode, and screen-print gold electrode (C220 AT, Dropsens®). The polymeric nanocomposites were characterized by electrochemical techniques, FTIR, SEM, elemental mapping, XPS, surface free energy, and contact angle. The concentration parameters of the polymeric solutions incorporated nanoparticle mass and the immobilization of anti-Tg antibodies on the modified electrodes were characterized by cyclic voltammetry. Incubation and interference assays were performed, as well as optimized anti-Tg, Tg deposition time, and incubation time using square wave voltammetry. Analytical curves were constructed from PVDF-NP TiO2-MP, PVDF-NT TiO2-MP, PVDF-OxG-MP, and PVDF-NP Au-MP films relating the suppression of the redox probe ioxi signal as a function of Tg concentration. R2 values, detection limit, quantitation limit, and recovery factor were obtained and compared with Roche® reference values. The streptavidin-biotin system attributed to the obtained electrochemical immunosensors optimum sensitivity, specificity, stability, and reproducibility. The streptavidin-biotin system attributed to the obtained electrochemical immunosensors optimum sensitivity, specificity, stability, and reproducibility. The proposed immunosensors were able to determine Tg at the ultrasensitive level, reaching values comparable to the LD of 0.04 ng/mL obtained by Roche®. Immunosensors have great potential to monitor Tg levels in patients undergoing total thyroidectomy and radioactive iodine ablation to screen for CDT metastasis.