scholarly journals Isolation of Extracellular Vesicles from Cell Culture Media by Differential Ultracentrifugation  v1 (protocols.io.bnr3md8n)

protocols.io ◽  
2020 ◽  
Author(s):  
Dima Ter ◽  
Wendy Trieu ◽  
Maia Norman ◽  
Roey Lazarovits ◽  
George Church ◽  
...  
Keyword(s):  
Cell Culture ◽  
Extracellular Vesicles ◽  
Culture Media ◽  
Cell Culture Media

Isolation and characterization of extracellular vesicles

2019 ◽  
Vol 1 (1) ◽  
pp. 18-26
Author(s):  
Fabia Fricke ◽  
Dominik Buschmann ◽  
Michael W. Pfaffl
Keyword(s):  
Cell Culture ◽  
Extracellular Vesicles ◽  
Culture Media ◽  
Body Fluids ◽  
Cell Culture Media ◽  
Advantages And Disadvantages ◽  
The Past ◽  
Isolation And Characterization ◽  
Basic Biology

Research into extracellular vesicles (EVs) gained significant traction in the past decade and EVs have been investigated in a wide variety of studies ranging from basic biology to diagnostic and therapeutic applications. Since EVs are secreted by most, if not all, eukaryotic and prokaryotic cells, they have been detected in body fluids as diverse as blood, urine and saliva as well as in cell culture media. In this chapter, we will provide an overview of EV isolation and characterization strategies and highlight their advantages and disadvantages.

scholarly journals Diffusion-Based Separation of Extracellular Vesicles by Nanoporous Membrane Chip

Biosensors ◽  
2021 ◽  
Vol 11 (9) ◽  
pp. 347
Author(s):  
Gijung Kim ◽  
Min Chul Park ◽  
Seonae Jang ◽  
Daeyoung Han ◽  
Hojun Kim ◽  
...  
Keyword(s):  
Cell Culture ◽  
Human Serum ◽  
Extracellular Vesicles ◽  
Culture Media ◽  
High Yield ◽  
Polycarbonate Membrane ◽  
Cell Culture Media ◽  
Nanoporous Membrane ◽  
Selective Filter ◽  
Novel Biomarkers

Extracellular vesicles (EVs) have emerged as novel biomarkers and therapeutic material. However, the small size (~200 nm) of EVs makes efficient separation challenging. Here, a physical/chemical stress-free separation of EVs based on diffusion through a nanoporous membrane chip is presented. A polycarbonate membrane with 200 nm pores, positioned between two chambers, functions as the size-selective filter. Using the chip, EVs from cell culture media and human serum were separated. The separated EVs were analyzed by nanoparticle tracking analysis (NTA), scanning electron microscopy, and immunoblotting. The experimental results proved the selective separation of EVs in cell culture media and human serum. Moreover, the diffusion-based separation showed a high yield of EVs in human serum compared to ultracentrifuge-based separation. The EV recovery rate analyzed from NTA data was 42% for cell culture media samples. We expect the developed method to be a potential tool for EV separation for diagnosis and therapy because it does not require complicated processes such as immune, chemical reaction, and external force and is scalable by increasing the nanoporous membrane size.

scholarly journals Isolation of small extracellular vesicles from cell culture supernatant

2020 ◽  
Author(s):  
Eun-Ju Im ◽  
Chan-Hyeong Lee ◽  
Moon-Chang Baek
Keyword(s):  
Cell Culture ◽  
Extracellular Vesicles ◽  
Culture Supernatant ◽  
Culture Media ◽  
Cell Culture Supernatant ◽  
Cell Culture Media

Abstract This protocol describes how to isolate small extracellular vesicles (sEV) from cell culture media using an ultracentrifuge and analyze the number of sEV by nanoparticle tracking analyzer.

Cell culture media: NMR approaches to evaluating quality and nutrient consumption

Planta Medica ◽  
2016 ◽  
Vol 81 (S 01) ◽  
pp. S1-S381
Author(s):  
KB Killday ◽  
AS Freund ◽  
C Fischer ◽  
KL Colson
Keyword(s):  
Cell Culture ◽  
Culture Media ◽  
Cell Culture Media ◽  
Nutrient Consumption

Separation of Extracellular Vesicles by DNA-Directed Immunocapturing Followed by Enzymatic Release

2020 ◽  
Author(s):  
Dario Brambilla ◽  
Laura Sola ◽  
Elisa Chiodi ◽  
Natasa Zarovni ◽  
Diogo Fortunato ◽  
...  
Keyword(s):  
Cell Culture ◽  
Extracellular Vesicles ◽  
Culture Media ◽  
Biological Fluids ◽  
Imaging Techniques ◽  
Cell Communication ◽  
Disease Diagnosis ◽  
Cell Culture Supernatant ◽  
Transmission Electron ◽  
Downstream Analysis

Extracellular vesicles (EVs) have attracted great interest among researchers due to their role in cell-cell communication, disease diagnosis, and drug delivery. In spite of their potential in the medical field, there is no consensus on the best method for separating microvesicles from cell culture supernatant and complex biological fluids. Obtaining a good recovery yield and preserving physical characteristics is critical for the diagnostic and therapeutic use of EVs. The separation is made complex by the fact that blood and cell culture media, contain a large number of nanoparticles in the same size range. Methods that exploit immunoaffinity capture provide high purity samples and overcome the issues of currently used separation methods. However, the release of captured nanovesicles requires harsh conditions that hinder their use in certain types of downstream analysis. Herein, a novel capture and release approach for small extracellular vesicles (sEVs), based on DNAdirected immobilization of antiCD63 antibody is presented. The flexible DNAlinker increases the capture efficiency and allows releasing of EVs by exploiting the endonucleasic activity of DNAse I. This separation protocol works under mild conditions, enabling the release of intact vesicles that can be successfully analyzed by imaging techniques. In this article sEVs recovered from plasma were characterized by established techniques for EVs analysis including nanoparticle tracking and transmission electron microscopy.<br>

scholarly journals Immunomagnetic sequential ultrafiltration (iSUF) platform for enrichment and purification of extracellular vesicles from biofluids

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jingjing Zhang ◽  
Luong T. H. Nguyen ◽  
Richard Hickey ◽  
Nicole Walters ◽  
Xinyu Wang ◽  
...  
Keyword(s):  
Extracellular Vesicles ◽  
Culture Media ◽  
Metastatic Breast ◽  
Clinical Samples ◽  
Clinical Use ◽  
Immunomagnetic Beads ◽  
Liquid Biopsies ◽  
Cell Culture Media ◽  
Non Invasive ◽  
Healthy Donors

AbstractExtracellular vesicles (EVs) derived from tumor cells have the potential to provide a much-needed source of non-invasive molecular biomarkers for liquid biopsies. However, current methods for EV isolation have limited specificity towards tumor-derived EVs that limit their clinical use. Here, we present an approach called immunomagnetic sequential ultrafiltration (iSUF) that consists of sequential stages of purification and enrichment of EVs in approximately 2 h. In iSUF, EVs present in different volumes of biofluids (0.5–100 mL) can be significantly enriched (up to 1000 times), with up to 99% removal of contaminating proteins (e.g., albumin). The EV recovery rate by iSUF for cell culture media (CCM), serum, and urine corresponded to 98.0% ± 3.6%, 96.0% ± 2.0% and 94.0% ± 1.9%, respectively (p > 0.05). The final step of iSUF enables the separation of tumor-specific EVs by incorporating immunomagnetic beads to target EV subpopulations. Serum from a cohort of clinical samples from metastatic breast cancer (BC) patients and healthy donors were processed by the iSUF platform and the isolated EVs from patients showed significantly higher expression levels of BC biomarkers (i.e., HER2, CD24, and miR21).

Selection and preliminary application of DNA aptamer targeting A549 excreta in cell culture media

2021 ◽  
pp. 106811
Author(s):  
Yuanbin Guo ◽  
Ming Shi ◽  
Xiujuan Liu ◽  
Huagang Liang ◽  
Liming Gao ◽  
...  
Keyword(s):  
Cell Culture ◽  
Culture Media ◽  
Dna Aptamer ◽  
Cell Culture Media ◽  
Preliminary Application
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