Engineering nanozymes with robust chemiluminescence activity for rapid and quantitative detection of Epstein-Barr...

The strong correlation between Epstein-Barr virus (EBV) infection and diseases has been well established. However, there is still a lack of convenient methods for quantitative detection of EBV DNA. Here, we synthesize an iron-based single-atom nanozyme (MFePH) with high chemiluminescence activity and integrate it with recombinase polymerase amplification (RPA) and Fe₃O₄-based magnetic separation for the sensitive and noninvasive detection of EBV DNA. Specifically, MFePH is synthesized by first coordinating histidine axially to the iron-porphyrin, thereby simulating the structure-activity relationship of the active site in natural peroxidase, followed by its integration into a Zn-based metal-organic framework. MFePH demonstrates comparable chemiluminescence activity to that of natural peroxidase. In the RPA reaction, EBV DNA is amplified using a primer pair labelled with biotin and digoxin. The amplified products are enriched via anti-digoxin antibody-labelled magnetic Fe₃O₄. Concurrently, the streptavidin-labelled MFePH is responsible for signal amplification. This detection system accurately detects as low as 100 copies per mL of EBV DNA within 25 min, which is much shorter than the 2-3 h required by traditional qPCR (quantitative real-time polymerase chain reaction). Consequently, this study provides a convenient strategy for EBV DNA detection, which is expected to markedly facilitate early screening for EBV-associated diseases.