01. ExoSense: a microprobe-based method for single-step isolation and genetic of exosomes

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The goal of this study is to develop a new method for non-invasive and selective isolation of exosomes from cell media. Exosomes, a type of extracellular vesicles, contain proteins and RNA biomarkers for the diagnosis of diseases. The core of this technology is a stainless-steel microprobe (300μm × 30mm) functionalized with anti-CD63 antibodies that specifically capture pure exosomal subpopulations. This method provides several advantages over commercial exosome-purification technologies. These include increased selectivity, purification of an antigen-specific subpopulation of exosomes, and direct integration of the microprobe with genetic analysis instruments. Experiments were performed to assess the efficiency of the functionalization of the probe as well as the number of captured exosomes, exosomal protein, and RNAs per probe. Scanning electron imaging was used to visualize the polyelectrolyte coverage while fluorescent imaging was applied to determine the efficiency of the chemical bond of NH2-conjugated biotin to the COOH group of polyacrylic acid. The captured exosomes were quantified by measuring the activity of acetylcholinesterase (AChE). The BCA protein assay was utilized to quantify the amount of protein captured per microprobe while Agilent Bioanalyzer 2100 was used to assess the quantity and quality of the exosomal RNA. Our results indicate that the microprobe-based technology isolates CD63-positive exosomal subpopulation (23×106 exosomes/probe) from astrocytes derived cell media after 16 hours of incubation. The exosomal fraction is enriched in small RNAs. Future work will focus on the assessment of blocking reagents (BSA, PEG) to reduce the binding of the cell-media-derived protein to the surface of the probe.