Construction of Aptamer-Based Nanobiosensor for Breast Cancer Biomarkers Detection Utilizing g-C3N4/Magnetic Nano-Structure

Abstract

An electrochemical aptasensor has been developed to determine breast cancer biomarkers (CA 15-3). Aptamer chains were immobilized on the surface of the electrode by g-C₃N₄/Fe₃O₄ nanoparticles, which increased the conductivity and active surface area of the electrode. X-ray diffraction analysis (XRD), Fourier-transformed infrared spectroscopy (FTIR), and transmission electron microscopy (TEM) measurements have been carried out to characterize the nanomaterials. Cyclic voltammetry, square wave voltammetry, and electrochemical impedance spectroscopy have been used to characterize the developed electrode. The results demonstrate that the modified electrode has better selectivity for CA 15-3 compared to other biological molecules. It has a good electrochemical response to CA 15-3 with a detection limit of 0.2 UmL^-1 and a linear response between 1 and 9 UmL^-1. It has been used as a label-free sensor in potassium ferrocyanide medium and as methylene blue-labeled in phosphate buffer medium. This electrode was successfully applied to analyze the serum of diseased and healthy individuals, which corroborates its high potential for biosensing applications, especially for the diagnosis of breast cancer.

Keywords: aptamer; biomarker; biosensor; breast cancer; g-C3N4; nanomaterial.

Figure 1

(a) X-ray pattern of g-C₃N₄ and g-C₃N₄/Fe₃O₄. (b) FTIR spectra of g-C₃N₄ and g-C₃N₄/Fe₃O₄ and g-C₃N₄/Fe₃O₄/Apt. (c) TEM image of g-C₃N₄ and g-C₃N₄/Fe₃O_4.

Figure 2

Electrochemical characterization of the developed nanobiosensor by (a) CV analysis, (b) SWV analysis and (c) EIS analysis of the bare electrode and each step of the preparation of the modified electrodes in $K_4\left[Fe{\left(CN\right)}_6\right]{ }^{\raisebox{1ex}{$-3$}\!\left/ \!\raisebox{-1ex}{$-4$}\right.}$ (0.2 mM).

Figure 3

Time profile of aptamer/nanobiosensor interaction based on SWV techniques at different incubation times.

Figure 4

(a) Stability test of the electrode labeled by methylene blue. (b) Percentage of electrochemical stability of methylene blue electrode.

Figure 5

(a) CV analysis, (b) SWV voltammograms, and (c) Nyquist diagrams of the electrode in $K_4\left[Fe{\left(CN\right)}_6\right]{ }^{\raisebox{1ex}{$-3$}\!\left/ \!\raisebox{-1ex}{$-4$}\right.}$ (0.2 mM) media at different concentrations of CA 15-3 ranging from 0 to 90 UmL⁻¹. (d,e) CV and SWV linear responses of the nanoprobe to different concentrations of CA 15-3.

Figure 6

(a) CV and (b) SWV test results at (c) various concentrations of CA 15-3 with the labeled electrode by methylene blue.

Figure 7

(a) Diffusion control test of the electrode labeled by methylene blue. (b) Calibration curve of diffusion control test of the electrode labeled by methylene blue in the reduction path. (c) Calibration curve of diffusion control test of the electrode labeled by methylene blue in the oxidation path.

Figure 8

Comparison of nanoprobe electrode signal for CA 15-3 (5 UmL⁻¹) glucose (90 mg dL⁻¹), PSA (2.5 pg mL⁻¹), FBS (5 ng mL⁻¹), BSA (7 μg mL⁻¹).

Figure 9

Real sample test results based on SWV techniques.