Point-of-care detection of prostate cancer through surface protein profiling on extracellular vesicles using colorimetric 3D paper-based analytical devices.

Surface proteins on tumor-derived extracellular vesicles (EVs) are frequently associated with tumorigenesis and progression. The effective evaluation of surface protein profiles on plasma EVs provides a reliable method for cancer diagnosis due to their high abundance (>108 particles/mL) and sustainable availability. However, quantifying the protein population on EVs is hampered by the difficulty of isolation, and the limited sensitivity of current EVs protein assays. In this study, we developed novel 3D paper-based analytical devices (PADs) for the digital quantification of EVs based on a colorimetric reaction. EVs were specifically captured by antibodies pre-stored on the paper, and the high affinity of Fe3+-doped DSPE-PDSNS for EVs membranes enabled signal amplification. The assay quantifies the protein level of EVs by measuring the intensity of the multicolor Prussian blue on the 3D PADs. The entire assay process on this platform is executed rapidly by folding and sliding the paper. This strategy achieves a detection limit of 3 × 103 particles/mL within half an hour, devoid of any reliance on instrument. Our work demonstrated that the EVs protein profiles (PSMA, PSA, PTK-7, EpCAM and EGFR) obtained using 3D PADs could successfully differentiate between healthy donors, prostatitis patients and prostate cancer (PCa) patients with an accuracy of 97.5 %. This versatile 3D PADs holds significant potential for the future development of point-of-care detection for cancer diagnosis in clinical setting.