Spray Modification of Amino-Functionalized g-C₃N₄ Nanosheets on Screen-Printed Carbon Electrodes for the Electrochemiluminescence Immunoassay of SARS-CoV-2 N Protein Based on Coreactant Release from Polymersome Tags

A signal-on electrochemiluminescence (ECL) immunoassay was developed for the detection of the SARS-CoV-2 N protein using coreactant-encapsulated polymersomes as tags and a graphitic carbon nitride (g-C3N4) nanosheet-modified screen-printed carbon electrode (SPCE) as the working electrode. Poly(ether imide) (PEI)-modified g-C3N4 nanosheets were prepared through ultrasonic exfoliation followed by a supramolecular covalent scaffold strategy. Spray coating, a scalable deposition technique, was employed to modify the SPCE surface with g-C3N4 nanosheets. After thermal curing, the nanosheets adhered firmly to the electrode via binders in the carbon ink, yielding a stable modified SPCE. Immunosensors were constructed by immobilizing capture antibodies on the g-C3N4-modified SPCE. Potassium persulfate, (K2S2O8), the coreactant of g-C3N4, was encapsulated in polymersomes to prepare ECL tags, and probes were fabricated by conjugating SARS-CoV-2 N protein binding aptamers to these polymersomes. Upon recognition of the target protein and sandwich complexes formed on the immunosensor, the subsequent addition of Triton X-100 disrupted the polymersomes, releasing large amounts of K2S2O8, which markedly enhanced the ECL mission of g-C3N4 nanosheets. This amplification enabled highly sensitive detection of the SARS-CoV-2 N protein. These findings demonstrate that controlled release of encapsulated coreactants from polymersome tags is an effective strategy to trigger g-C3N4-based ECL emission for sensitive viral protein detection.