Abstract: Methods and systems for characterizing extracellular vesicular biomarkers using a biochip with gold nanoparticles. The biochip includes a glass surface, a gold film layer on the glass surface, a plurality of gold nanoparticles coupled to the gold film layer, and a plurality of biotinylated antibodies coupled to the gold nanoparticles. In some implementations, the gold film layer of the biochip is coated with polyethylene glycol (PEG). The biotinylated antibodies are selected to capture specific types of extracellular vesicles. PD-L1/PD-1 proteins and RNAs in extracellular vesicles were characterized for cancer immunotherapy.

Abstract: Methods for purifying and isolating extracellular vesicles (EVs) from a biofluid using a sequential processing. Tangential flow filtration is applied to the biofluid to increase the concentration of EVs in the biofluid. After this is achieved, enrichment mode is halted and the biofluid is processed in diafiltration mode to remove contaminants (up to 99.9%). After performing the tangential flow filtration step, the concentration of EVs in the biofluid is further increased by ultracentrifugal filtration. After performing the ultracentrifugal filtration step, EVs of a particular target type are separated from other EVs by immunomagnetic affinity separation. In some implementations, the methods are used to isolate and quantify tumor EVs for cancer evaluation. Additionally, these methods can be used with a scaling factor to quantify EVs from a less concentrated biofluid such as, for example, urine.