Rapid and sensitive detection of cancer-derived small extracellular vesicles using Janus particles

Detecting small extracellular vesicles is critical for understanding disease biology and developing diagnostic tools, yet current methods require lengthy isolation steps and lack sensitivity owing to interference from abundant proteins. Here we report on an assay that uses Janus particles that enable rapid, isolation-free detection by exploiting Brownian rotation-induced blinking changes. When vesicles bind, their size significantly alters the blinking frequency, while smaller proteins produce no signal, ensuring selectivity. Using less than 10 μl of sample, the assay detects approximately 200 vesicles per microlitre and works directly on plasma, serum, urine and cell media in under 1 h. In a blind study of 87 subjects with colorectal cancer, pancreatic ductal adenocarcinoma, glioblastoma, Alzheimer's disease and healthy controls, the method identified disease type with an area under the curve of 0.90-0.99. Compared with ultracentrifugation combined with surface plasmon resonance, which requires 24 h, our approach delivers 2 orders of magnitude better sensitivity and dynamic range, offering a fast and robust platform for clinical and research applications.