Highly Effective Detection of Exosomal miRNAs in Plasma Using Liposome-Mediated Transfection CRISPR/Cas13a

Exosomal miRNAs play a critical role in cancer biology and could be potential biomarkers for cancer diagnosis. However, due to the low abundance of miRNAs in the exosomes, recognizing and detecting disease-associated exosomal miRNAs in an easy-to-operate way remain a challenge. Herein, we used a liposome-mediated membrane fusion strategy (MFS) to transfect CRISPR/Cas13a into exosomes, termed MFS-CRISPR, directly measuring exosomal miRNAs in plasma. Using the MFS-CRISPR platform for detection of the exosomal miR-21, we achieve a linear range spanning four orders of magnitude (10⁴-10⁸ particles/mL) and the method is able to detect the exosomal miR-21 in as low as 1.2 × 10³ particles/mL. The liposome-mediated MFS could confine fluorescent signals in fused vesicles, which can be used for exosome heterogeneity analysis. Moreover, MFS-CRISPR assay was evaluated by measuring clinical samples, and the difference of miR-21 expression of breast cancer patients and healthy donors was significant. Because of high sensitivity and simplicity, the proposed method could have promising clinical potential for cancer diagnosis and treatment monitoring.