Extracellular vesicles nanoarray technology: Immobilization of individual extracellular vesicles on nanopatterned polyethylene glycol-lipid conjugate brushes

• Published in: PloS one Vol. 14; no. 10; p. e0224091
• Main Authors: Yokota, Shusuke, Kuramochi, Hiromi, Okubo, Kyohei, Iwaya, Akiko, Tsuchiya, Shoichi, Ichiki, Takanori
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 24.10.2019; Public Library of Science (PLoS)
• Subjects: Aspect ratio; Atomic force microscopy; Biology and Life Sciences; Biomarkers; Biomaterials; Brushes; Cancer; Cardiovascular disease; Cell lines; Chemical modification; Conjugates; Coupling (molecular); Coupling agents; Cytology; Deformability; Density; Deoxyribonucleic acid; DNA; Electron beam lithography; Engineering and Technology; Engineering schools; Extracellular vesicles; Extracellular Vesicles - chemistry; Extracellular Vesicles - metabolism; Formability; Glycols (Class of compounds); HEK293 Cells; Humans; Immobilization; Lab-On-A-Chip Devices; Lipids; Lipids - chemistry; Membrane lipids; Membranes; Metastasis; Microfluidic devices; Microfluidics; MicroRNAs; Microscopy; Microscopy, Atomic Force; Morphology; Nanopatterning; Nanotechnology; Nanotechnology - instrumentation; Nanotechnology - methods; Organic chemistry; Particle size; Physical Sciences; Polyethylene glycol; Polyethylene Glycols - chemistry; Polyols; Production processes; Proteins; Proteomics; Research and Analysis Methods; Signal transduction; Silanes; Silicon; Studies; Technology; Tethering; Vesicles; Japan; New York; United States > US
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0224091

Arraying individual extracellular vesicles (EVs) on a chip is expected one of the promising approaches for investigating their inherent properties. In this study, we immobilized individual EVs on a surface using a nanopatterned tethering chip-based versatile platform. A microfluidic device was used to ensure soft, reproducible exposure of the EVs over the whole chip surface. The device is incorporated with a high-density nanoarray chip patterned with 200-nm diameter nanospots composed of polyethylene glycol (PEG)-lipid conjugate brushes. We present a procedure adopted for fabricating high-density PEG-lipid modified nanospots (200 nmϕ, 5.0 × 105 spots/mm2 in 2 × 2 mm2 area). This procedure involves nanopatterning using electron beam lithography, followed by multistep selective chemical modification. Aqueous treatment of a silane coupling agent, used as a linker between PEG-lipid molecules and the silicon surface, was the key step that enabled surface modification using a nanopatterned resist film as a mask. The nanoarray chip was removed from the device for subsequent measurements such as atomic force microscopy (AFM). We developed a prototype device and individually immobilized EVs derived from different cell lines (Sk-Br-3 and HEK293) on tethering nanospots. We characterized EV's morphology using AFM and showed the possibility of evaluating the deformability of EVs using the aspect ratio as an indicator.