Versatile bioluminescence CRISPR/Cas platform for point-of-care detection of biomarkers in whole blood

Clustered regularly interspaced short palindromic repeats (CRISPR) systems have achieved significant advancements in precise molecular diagnosis. However, their applications in whole blood detection remain challenging due to signal interference from blood autofluorescence. Here, we proposed a universal and accessible bioluminescent CRISPR/Cas (bioLUCas) platform for direct detection of disease biomarkers in whole blood. By employing a specially designed cpHNLuc-MB reporter, the bioLUCas system converts CRISPR/Cas12a trans-cleavage activity into a ratiometric bioluminescent signal, producing a distinct emission color change. Compared to conventional CRISPR/Cas12a-based sensors, this platform eliminates the need for external light excitation, effectively bypassing blood autofluorescence and offering high sensitivity. Additionally, the visual signal of bioLUCas system allows user-friendly readout methods, such as smartphone. The platform successfully facilitated point-of-care test (POCT) for myeloperoxidase (MPO) in clinical acute myelogenous leukemia (AML) blood samples and hepatitis C virus (HCV) RNA in synthetic blood samples. This work may advance CRISPR/Cas technology for accessible whole-blood disease diagnostics.