In situ detection of bacteria from skin interstitial fluid via CRISPR microneedles: An amplification-free platform...

We report a CRISPR/Cas12a-functionalized microneedle (MN) biosensor for in situ, amplification-free detection of pathogenic bacteria directly from skin interstitial fluid. The platform integrates conductive poly(styrene)/gold/graphene oxide MNs with a minimized electrochemical signal transducer, enabling real-time and highly specific sensing of bacterial DNA through CRISPR-mediated cleavage of ferrocene-labeled ssDNA reporters. Using Staphylococcus aureus as a model target, this strategy achieves direct detection limits of 0.69 pM in ex situ assays and 6.3 pM in situ, without the need for target amplification. The sensor also detects bacterial loads as low as 4.27 × 10⁵ CFU/mL in interstitial fluid, which is below typical clinical thresholds. Engineered for biocompatibility and minimal invasiveness, the MNs effectively access interstitial fluid, capture targets and produce quantifiable electrochemical signals within 1 h. Paired with a portable reader and smartphone interface, this wearable system offers a user-friendly tool for point-of-care diagnostics and personalized infectious disease monitoring.