Loop-mediated isothermal amplification combined with lateral flow biosensor for rapid and sensitive detection of monkeypox virus.

Abstract

The ongoing outbreak of the monkeypox, caused by monkeypox virus (MPXV), has been a public health emergency of international concern, indicating an urgent need for rapid and sensitive MPXV detection. Here, we designed a diagnostic test based on loop-mediated isothermal amplification (LAMP) and nanoparticle-based lateral flow biosensor(LFB)for diagnosis of MPXV infection, termed MPX-LAMP-LFB. A set of six LAMP primers was designed based the ATI gene of MPXV, and LAMP amplification of MPXV templates was performed at 63&#xb0;C for only 40&#x2009;min. The results were rapidly and visually decided using the LFB test within 2&#x2009;min. The MPX-LAMP-LFB assay can specifically detect MPXV strains without cross-reaction with non-MPXV pathogens. The sensitivity of the MPX-LAMP-LFB assay is as low as 5 copies/&#x3bc;l of plasmid template and 12.5 copies/&#x3bc;l of pseudovirus in human blood samples. The whole process of the MPX-LAMP-LFB assay could be completed ~1&#x2009;h, including rapid template preparation (15&#x2009;min), LAMP reaction (40&#x2009;min)and result reporting (<2&#x2009;min). Collectively, MPX-LAMP-LFB assay developed here is a useful tool for rapid and reliable diagnosis of MPXV infection.