We and others have previously shown that the UC step damages the vesicles and leads to aggregation 18, 19, 20, which can ultimately affect downstream analysis 21 or application of EVs 19, 22. The gold standard in the field is to purify EVs by sequential centrifugation followed by an ultracentrifuge (UC) step to pellet the EVs at 110,000 × g 18. EVs are of fundamental importance in conveying critical intercellular messages 8, 10 both in physiological and pathological processes, such as taking part in the coagulation cascade 11, immune response 12, 13, 14 as well as aiding the spread of malignancies 9, 15 and viral infections 16, 17.īecause of their small size, physicochemical properties and the complexity of the surrounding fluid, purification of EVs is a great challenge. They carry proteins and RNAs, both miRNAs and mRNAs, and have been shown to transfer their cargo to recipient cells 3, 8, 9. Exosomes are 70–150 nm in size and originate from the endocytic pathway 5 whereas MVs are generally larger, 100–1000 nm in diameter and bud directly from the plasma membrane 6, 7. In this article, the term EVs will refer to exosomes and MVs only. Hence, the BE-SEC based EV purification method represents an important methodological advance likely to facilitate robust and reproducible studies of EV biology and therapeutic application.Įxtracellular vesicles (EVs) are nanosized cell-derived vesicles 1, 2, 3 delimited by a lipid bilayer and typically divided into three subgroups, according to their biogenesis pathways exosomes, microvesicles (MVs) and apoptotic bodies 4. Furthermore, uptake of eGFP labelled EVs in recipient cells was comparable between BE-SEC and UC samples. This technique is reproducible and scalable, and surface marker analysis by bead-based flow cytometry revealed highly similar expression signatures compared with UC-purified samples. Here we show that commercially available bind-elute size exclusion chromatography (BE-SEC) columns purify EVs with high yield (recovery ~ 80%) in a time-efficient manner compared to current methodologies. EVs have traditionally been purified by ultracentrifugation (UC), however UC has limitations, including resulting in, operator-dependant yields, EV aggregation and altered EV morphology, and moreover is time consuming. The pores of the bead outer layer prevent large targets (M r 700 000) from binding to the ligands while smaller impurities can enter freely into the beads where they are captured.Extracellular vesicles (EVs) play a pivotal role in cell-to-cell communication and have been shown to take part in several physiological and pathological processes. These internalized ligands bind impurities strongly over a wide range of pH and salt concentration. The octylamine ligands found in the core of the beads are multimodal, being both hydrophobic and positively charged. Capto Core 700 is a novel, layered bead chromatography medium consisting of a porous outer layer and ligand-activated core. Prepacked HiTrap columns for easy screening and convenient process development, as well as small scale purification. Wide operational window of pH and conductivity. High capacity and productivity in flowthrough mode. HiTrap Capto Core 700 is a ready-to-use 1 ml column, prepacked with Capto Core 700, for intermediate purification and polishing of viruses and other large biomolecules in flow-through mode, Each bead has a ligand-activated core and an outer layer without ligands, The layer prevents the large targets from entering into the beads whereas proteins and other impurities enter into the core where they bind to the hydrophobic and positively charged octylamine ligands, HiTrap columns are easy used manually together with a syringe, with a lab pump or a chromatogtaphy system such as ÄKTA design, HiTrap Capto Core 700 HiTrap Capto Core 700 is prepacked with Capto Core 700, a multimodal BioProcess medium for intermediate purification and polishing of viruses and other large biomolecules. HiTrap Capto Core 700, 5 x 1 mlHiTrap Capto Core 700, 5 x 1 ml
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