A nanohybrid composed of Prussian Blue and graphitic C3N4 nanosheets as the signal-generating tag in an enzyme-free electrochemical immunoassay for the neutrophil gelatinase-associated lipocalin

An enzyme-free electrochemical immunoassay is described for the neutrophil gelatinase-associated lipocalin (NGAL; a biomarker of kidney disease). Prussian Blue (PB) nanoparticles with redox activity were deposited on graphitic C3N4 nanosheets (g-C3N4) by in-situ reduction. A screen printed electrode (SPCE) was modified with antibody against NGAL, and the PB-g-C3N4 nanohybrid was used as the signal-generating tag for the secondary antibody against NGAL. Upon addition of target NGAL and of secondary antibody, a sandwich is formed on the SPCE. At an applied potential of typically 0.13 V (vs. Ag/AgCl), a well-defined voltammetric peak is observed that results from the presence of PB on the secondary antibody. Under optimal conditions, the peak current increases linearly in the 0.01 to 10 ng.mL(-1) NGAL concentration range, and the detection limit is 2.8 pg.mL(-1). An average precision of < 12% was accomplished in the batch-to-batch mode. Other disease-related biomarkers do not interfere. The accuracy and inter-laboratory validation of this method were evaluated for target NGAL detection in spiked human serum by using a commercial ELISA. The results obtained by the two methods are in good accordance.