Sustainable and portable CRISPR-based diagnostics for high-sensitivity Mpox detection

Monkeypox (Mpox) has emerged as a critical public health challenge, creating an urgent need for rapid, reliable, field-deployable diagnostic tools for outbreak scenarios. Here, we present Kairo-CONAN, a novel CRISPR-Cas3-based point-of-care diagnostic platform for Mpox, engineered for sustainability and portability. This system uses a disposable hand warmer as a stable heat source and incorporates freeze-dried reagents for ambient temperature stability, enabling device-free, sensitive detection by lateral flow assay strips. Utilizing the unique DNA-targeting and cleavage properties of CRISPR-Cas3, we optimized probe DNA configurations for high specificity and designed clade-specific target CRISPR RNAs (crRNAs). Kairo-CONAN has demonstrated rapid, high-sensitivity, specific detection of Mpox virus (MPXV) DNA across multiple clades, including Clade Ia (Congo), Clade Ib (synthetic DNA), and Clade IIb (Tokyo). Kairo-CONAN addresses logistical and environmental challenges to offer a sustainable, cost-effective, field-adapted solution for infectious disease diagnostics, aligning with the 100 Days Mission framework to enhance global outbreak response efforts.