A novel electrochemical nanobiosensor for the ultrasensitive and specific detection of femtomolar-level gastric cancer biomarker miRNA-106a

Abstract

Gastric cancer (GC) is the second leading cause of cancer-related deaths all over the world. miR-106a is a circulatory oncogenic microRNA (miRNA), which overexpresses in various malignancies, especially in GC. In this study, an ultrasensitive electrochemical nanobiosensor was developed for the detection of miR-106a using a double-specific probe methodology and a gold-magnetic nanocomposite as tracing tag. The successful modification of the electrode and hybridization with the target miRNA were confirmed by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV) methods. Differential pulse voltammetry (DPV) was used for quantitative evaluation of miR-106a via recording the reduction peak current of gold nanoparticles. The electrochemical signal had a linear relationship with the concentration of the target miRNA ranging from 1 × 10^-3 pM to 1 × 10³ pM, and the detection limit was 3 × 10^-4 pM. The proposed miRNA-nanobiosensor showed remarkable selectivity, high specificity, agreeable storage stability, and great performance in real sample investigation with no pretreatment or amplification. Consequently, our biosensing strategy offers such a promising application to be used for clinical early detection of GC and additionally the screen of any miRNA sequence.

Keywords: electrochemical nanobiosensor; gastric cancer; gold–magnetic nanoparticle; miR-106a.

Figure 1

Schematic of the principal mechanism for miR-106a detection by the nanobiosensor. (1) Preparation the nanoprobe; (2) modification of the electrode; and (3) hybridization steps (TMC = N-trimethylchitosan).

Figure 2

(A) TEM images of synthesized Fe₃O₄ NPs (a), gold NPs (b), TMC@Fe₃O₄ NPs (c), and gold–magnetic NPs (d) with their corresponding particle size distribution (inset). (B) UV–vis analysis of gold NPs (a), Fe₃O₄ NPs (b), TMC@Fe₃O₄ NPs (c), and gold–magnetic NPs (d). (C) EDXD spectra of gold–magnetic NPs.

Figure 3

(A) AFM images of (a) a bare SPCE, (b) a SPCE after coating with streptavidin, (c) after immobilization of P2, and (d) after hybridization with the target complex. (B) Cyclic voltammograms carried out in 5.0 mM [Fe(CN)₆]^3−/4− solution containing 1.0 M KCl, at a scan rate of 100 mV/s for bare SPCE (blue), streptavidin-coated SPCE (purple), P2/streptavidin-coated SPCE (orange), and after hybridization with the target complex (green).

Figure 4

(A) Differential pulse voltammograms for the electrochemical detection of miR-106a upon serial dilutions of target miR-106a at scan rate 100 mV/s in 1 M HCl. The concentrations of target miRNA are: 0, 0.001, 0.01, 0.1, 1, 5, 10, 50, 100, 500, and 1000 pM. (B) The calibration curve of miR-106a as the relationship between current and logarithm of miR-106a concentration. Each data point is the average of five replicates.

Figure 5

Differences in signal intensities in presence of interference miR-15a (nc1), miR-21 (nc2), and miR-200c (nc3). (A) miR-106a (10 pM); (B) miR-106a (10 pM) + nc1 (10 pM) + nc2 (10 pM) + nc3 (10 pM); (C) nc1 (10 pM); (D) nc1 (50 pM); (E) miR-106a (10 pM) + nc1 (10 pM); (F) miR-106a (10 pM) + nc1 (50 pM); (G) nc2 (10 pM); (H) nc2 (50 pM); (I) miR-106a (10 pM) + nc2 (10 pM); (J) miR-106a (10 pM) + nc2 (50 pM); (K) nc3 (10 pM); (L) nc3 (50 pM); (M) miR-106a (10 pM) + nc3 (10 pM); (N) miR-106a (10 pM) + nc3 (50 pM).