Fabrication of a Label-free Electrochemical Aptasensor to Detect Cytochrome C in the Early Stage of Cell Apoptosis

A label-free direct electrochemical aptasensor is presented for the identification of cytochrome c (Cyt c) at the nM concentration level. Carbon nanofibers (CNF), as a highly conductive material, were used to modify a glassy carbon electrode (GCE) and thus increase its conductivity. Moreover, to enhance the immobilization of aptamers (Apt) on the electrode surface, graphene oxide functionalized with aspartic acid (GOAsp) was added to the surface. Aspartic acid with countless carboxyl groups (-COOH) on its surface caused more aptamers to be immobilized on the electrode surface. Electrochemical impedance spectroscopy (EIS), cyclic voltammetry (CV), and differential pulse voltammetry (DPV) were used to monitor the step-by-step fabrication of the label-free direct electrochemical aptasensor. The label-free quantification of Cyt c was also done by the direct electron transfer between the Fe(III)/Fe(II)-heme redox-active sites which were selectively bound to the aptamers on the GCE and the surface of the electrode. Under optimum conditions, the peak currents of differential pulse voltammograms at 0.26 V (vs. Ag/AgCl) were used for calibration. The proposed aptasensor performs in a wide dynamic range from 10 nM to 100 µM with a low detection limit of 0.74 nM for cytochrome c. It also has high selectivity as well as acceptable stability. These advantages make the biosensor capable of detecting early-stage apoptotic cells that contribute to early cancer diagnosis.