Novel electrochemical platform based on C3N4-graphene composite for the detection of neuron-specific enolase as a biomarker for lung cancer

Abstract

Lung cancer remains the leading cause of cancer mortality worldwide. Small cell lung cancer (SCLC) accounts for 10-15% of cases and has an overall 5-years survival rate of only 15%. Neuron-specific enolase (NSE) has been identified as a useful biomarker for early SCLC diagnosis and therapeutic monitoring. This work reports an electrochemical immunosensing platform based on a graphene-graphitic carbon nitride (g-C₃N₄) nanocomposite for ultrasensitive NSE detection. The g-C₃N₄ nanosheets and graphene nanosheets were synthesized via liquid exfoliation and integrated through self-assembly to form the nanocomposite. This nanocomposite was used to modify screen-printed carbon electrodes followed by covalent immobilization of anti-NSE antibodies. The unique properties of the graphene-g-C₃N₄ composite facilitated efficient antibody loading while also enhancing electron transfer efficiency and electrochemical response. Systematic optimization of experimental parameters was performed. The immunosensor exhibited a wide linear detection range of 10 pg/mL to 100 ng/mL and low limit of detection of 3 pg/mL for NSE along with excellent selectivity against interferences. Real serum matrix analysis validated the applicability of the developed platform for sensitive and accurate NSE quantifica-tion at clinically relevant levels. This novel graphene-g-C₃N₄ nanocomposite based electro-chemical immunoassay demonstrates great promise for early diagnosis of SCLC.

Figure 1

Fabrication process of proposed electrochemical sensor.

Figure 2

SEM image of (A) g-C₃N₄ nanosheets and (B) graphene-g-C₃N₄ composite.

Figure 3

(A) XRD patterns of g-C₃N₄ nanosheets and graphene-g-C₃N₄ composite. (B) Raman spectra of g-C₃N₄ nanosheets, graphene and graphene-g-C₃N₄ composite.

Figure 4

FTIR spectra of g-C₃N₄ nanosheets, graphene and graphene-g-C₃N₄ composite.

Figure 5

Cyclic voltammograms of bare SPCE, graphene/SPCE, g-C₃N₄-graphene/SPCE, and anti-NSE/g-C₃N₄-graphene/SPCE in 0.1 M PBS containing 5 mM [Fe(CN)₆]^3−/4− at a scan rate of 50 mV/s.

Figure 6

Nyquist plots obtained from electrochemical impedance spectroscopy of bare SPCE, graphene/SPCE, g-C₃N₄-graphene/SPCE, and anti-NSE/g-C₃N₄-graphene/SPCE in 0.1 M PBS containing 5 mM [Fe(CN)₆]^3−/4−. Inset shows the equivalent circuit model used for fitting the EIS data.

Figure 7

Effect of (A) incubation time; (B) incubation temperature; and (C) pH on the response of the immunosensor.

Figure 8

(A) Calibration curve of the immunosensor for different concentrations of NSE; (B) Selectivity of the immunosensor for 5 ng/mL NSE against other proteins.

Figure 9

(A) Correlation of determined and spiked NSE concentrations in serum samples; (B) Comparison of NSE levels in serum samples as measured by the immunosensor and ELISA.