Exosome purification based on PEG-coated Fe3O4 nanoparticles

Abstract

Cancer cells secrete many exosomes, which facilitate metastasis and the later growth of cancer. For early cancer diagnosis, the detection of exosomes is a crucial step. Exosomes exist in biological fluid, such as blood, which contains various proteins. It is necessary to remove the proteins in the biological fluid to avoid test interference. This paper presented a novel method for exosome isolation using Fe3O4 magnetic nanoparticles (MNPs), which were synthesized using the chemical co-precipitation method and then coated with polyethylene glycol (PEG). The experimental results showed that the diameter of the PEG-coated Fe3O4 nanoparticles was about 20 nm, while an agglomerate of MNPs reached hundreds of nanometers in size. In the protein removal experiments, fetal bovine serum (FBS) was adopted as the analyte for bioassays of exosome purification. PEG-coated Fe3O4 MNPs reduced the protein concentration in FBS to 39.89% of the original solution. By observing a particle size distribution of 30~200 nm (the size range of various exosomes), the exosome concentrations were kept the same before and after purification. In the gel electrophoresis experiments, the bands of CD63 (~53 kDa) and CD9 (~22 kDa) revealed that exosomes existed in FBS as well as in the purified solution. However, the bands of the serum albumins (~66 kDa) and the various immunoglobulins (around 160 ~ 188 kDa) in the purified solution’s lane explained that most proteins in FBS were removed by PEG-coated Fe3O4 MNPs. When purifying exosomes from serum, protein removal is critical for further exosome investigation. The proposed technique provides a simple and effective method to remove proteins in the serum using the PEG-coated Fe3O4 MNPs.

Comparative analysis of exosome isolation methods using culture supernatant for optimum yield, purity and downstream applications

“…We found variation (−2.8 mV to −29.12 mV) in the zeta potential of exosome preparations by different methods. Exosomes isolated by all the methods displayed negative magnitude of zeta potential similar to previous reports 30,31 . However, we observed the least negative value of exosomes isolated from Invitrogen precipitation method, which might be due to the presence of charged molecules that increased the ionic strength of the suspension medium (water).…”

Electrostatic Surface Properties of Blood and Semen Extracellular Vesicles: Implications of Sialylation and HIV-Induced Changes on EV Internalization

“…As in cells, the electrochemical property of the EV surfaces also depends on the outer membrane cargo, composed of an intermolecular network of lipids [ 13 ], proteins [ 14 ], glycans [ 15 ], and possibly nucleic acids [ 16 ]. While EV surface charge has scarcely been reported as ζ-potential values [ 17 , 18 , 19 , 20 ], ζ-potential alone in the absence of pH at which measurements were taken is insufficient to infer EV surface charge status. This is because pH variation drastically affects the surface charge [ 21 ].…”

Exosomes in Gliomas: Biogenesis, Isolation, and Preliminary Applications in Nanomedicine

“…A recent study from Chang et al [ 70 ] presented a novel method for exosome isolation using Fe 3 O 4 magnetic nanoparticles (MNPs) coated with polyethylene glycol (PEG). PEG chains can form reticular structures that allow the entrapping of proteins, aggregates, and impurities in the holes of MNP.…”