Desenvolvimento de método eletroanalítico utilizando sensores eletroquímicos descartáveis para detecção do biomarcador miRNA-203 visando o diagnóstico de câncer de cabeça e pescoço

Abstract

A great effort has been made in the development of new analytical methods for biomarker detection aiming at the diagnosis of cancer. Therefore, in this work it was developed an electroanalytical method using low-cost disposable electrochemical sensors (DESs) for the early diagnosis of head and neck cancer (HNC) in a simple, fast, and accurate way. The DESs were fabricated using a craft cutter printer and disposable materials, such as polyester sheets (transparency film for laser printers), vinyl adhesives, laminating pouch masks, carbon conductive and Ag/AgCl inks. All electrodes present in the DES were fabricated employing the screen printing technique, which were used for the detection of the microRNA-203 biomarker (miRNA-203) related to the HNC. The analytical method for the miRNA-203 detection was developed using a capture probe DNA sequence (DNACp) which was immobilized in magnetic particles (MPs) to capture miRNA-203 (MPCp) through hybridization assays resulting in the formation of the bioconjugate MPCp-miRNA-203, after the capture of miRNA-203 a detection probe DNA sequence (DNADp) complementary to DNACp was immobilized on gold nanoparticles (AuNPsDp) allowing its binding in the terminal region of DNACp forming the bioconjugate MPCp-miRNA203-AuNPsDp. The bioconjugate formed was characterized using the techniques Scanning Electron Microscopy, Transmission Electron Microscopy and, Energy Dispersive X-Ray Spectroscopy. The analytical signal was obtained using the Square Wave Voltammetry (SWV) technique and a 0.2 mol L−1 HCl solution as a supporting electrolyte. After the optimization of the SWV parameters, the method was applied in samples of total RNA from cell lines and lymph nodes of patients with HNC provided by the Molecular Oncology Research Center of the Barretos Cancer Hospital. The electroanalytical method developed presented an analytical curve with a linear range from 1.0 to 500.0 fmol L−1, linear correlation coefficient of 0.997, and limit of detection of 0.52 fmol L−1. The proposed method differentiated healthy individuals and patients with HNC through the amount of miRNA-203 found in the lymph node samples and was presented as an alternative method of low cost, very attractive for the early diagnosis of HNC in a simple, fast and, precise way, which could contribute to a more effective treatment to patients with the disease.