A ratiometric electrochemiluminescence method using a single luminophore of porous g-C3N4 for the ultrasensitive determination of alpha fetoprotein

Abstract

In this work, we report a simple ratiometric electrochemiluminescence (ECL) method for ultra-sensitive immunoanalysis. A glassy carbon electrode (GCE) was modified by a mixture of porous g-C₃N₄ nanosheets and carbon nanotubes (CNTs). Secondary antibodies were labeled using CuS nanoparticles as the tags. After immune recognition, CuS nanoparticles in the immunocomplex were dissolved as Cu²⁺, which can quench the ECL of g-C₃N₄. The amount of Cu²⁺ was determined to quantify the concentration of the target antigen. To enhance the sensitivity, Cu²⁺ ions were firstly enriched and reduced to Cu on the surface of GCE/CNTs-g-C₃N₄, and the cathodic ECL of g-C₃N₄ was measured as the reference signal in the ratiometric ECL measurements. After applying a potential of 0.6 V (vs. Ag/AgCl) for 6 s, Cu was dissolved as Cu²⁺, which can quench the ECL of g-C₃N₄ with much higher efficiency because the freshly dissolved Cu²⁺ ions were distributed mainly within the Helmholtz layer of GCE/CNTs-g-C₃N₄. By using the ECL intensity ratio of GCE/CNTs-g-C₃N₄ (Cu²⁺⁾ to GCE/CNTs-g-C₃N₄ (Cu) measured under the potentiostatic model as the signal indictor, the ratiometric ECL method was used to detect a biomarker of alpha fetoprotein with the limit of detection of 0.1 fg mL⁻¹. It was shown that the influence of the difference in electrode modification and ECL measurement conditions on the determination of Cu²⁺ is suppressed greatly in the ratiometric ECL method. The combination of ratiometric ECL with electrochemical enrichment and biometallization is a useful strategy to enhance the sensitivity and reproducibility in immunoanalysis.