High-precision phenotyping of breast cancer exosomes based on washable magnetic microarrays and super-resolution tricolor fluorescence co-localization

Exosome is a kind of membranous vesicles released from cells and carry a number of important signaling molecules, they play an important role in cellular communication, cell migration, angiogenesis as well as tumor cell growth. Exosome-based cancer diagnosis is usually achieved by detecting exosomal nucleic acids, lipids, and surface proteins, as they reflect tumor type and progression. Here, we proposed a method to rapidly prepare an array of washable magnetic nanoparticles (magnetic beads, MBs) by a magnetic field controlled system, which facilitate the analyzing of exosome phenotypes via super-resolution tricolor fluorescence co-localization (SRTFC) and pixel counting (CFPP). Firstly, nanopore arrays were designed and prepared by 3D printing technology. MBs@SiO2@Au nanospheres synthesized by hydrothermal method were rapidly absorbed into the nanopore arrays using a magnetic field to prepare a washable magnetic microarray substrate (WMMS). Then, exosomes were specifically labeled with three specific proteins to obtain the 3D phenotypic information of various exosomes. This method avoids meaningless and repetitive substrate preparation work and further improve the utility of SR-TFC, which is a high precision phenotyping strategy that we have recently proposed. This work provides a reliable and efficient exosome-based tumor detection platform, which is conducive to advancing the clinical application of SR-TFC.