scholarly journals Isolation of human salivary extracellular vesicles by iodixanol density gradient ultracentrifugation and their characterizations

2016 ◽  
Vol 5 (1) ◽  
pp. 30829 ◽  
Author(s):  
Kazuya Iwai ◽  
Tamiko Minamisawa ◽  
Kanako Suga ◽  
Yasutomo Yajima ◽  
Kiyotaka Shiba
Keyword(s):  
Density Gradient ◽  
Extracellular Vesicles ◽  
Density Gradient Ultracentrifugation

scholarly journals Comparative Evaluation of Methods for Isolating Small Extracellular Vesicles Derived from Pancreatic Cancer Cells

2021 ◽  
Author(s):  
Jie-Min Wang ◽  
Yong-Jiang Li ◽  
Jun-Yong Wu ◽  
Jia-Xin Cai ◽  
Jing Wen ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Density Gradient ◽  
Extracellular Vesicles ◽  
Pancreatic Cancer Cell Line ◽  
Size Exclusion ◽  
Density Gradient Ultracentrifugation ◽  
High Concentration ◽  
Protein Marker ◽  
Pancreatic Cancer Cells

Abstract Background: Small extracellular vesicles (sEVs) are nanosized vesicles involved in cell-to-cell communication. sEVs have been widely studied for clinical applications such as early detection of diseases and as therapeutics. Various methods for sEVs isolation are been using, but different methods may result in different qualities of sEVs and impact downstream analysis and applications. Here, we compared current isolation methods and performed a comparative analysis of sEVs from supernatant of cultured pancreatic cancer cells.Methods: Ultracentrifugation, ultrafiltration and co-precipitation as concentration methods were firstly evaluated for yield, size, morphology and protein level of pellets. Then, isolate sEVs obtained by four different purification methods: size exclusion chromatography, density gradient ultracentrifugation, ultracentrifugation, and immunoaffinity capturing, were analysed and compared.Results: For the concentration process, ultracentrifugation method obtained high quality and high concentration of pellets. For the purification process, immunoaffinity capturing method obtained the purest sEVs with less contaminants, while density gradient ultracentrifugation-based method obtained sEVs with the smallest size. Proteomic analysis revealed distinct protein contents of purified sEVs from different methods. Conclusions: For isolating sEVs derived from supernatant of cultured pancreatic cancer cell line, ultracentrifugation-based method is recommended for concentration of sEVs, density gradient ultracentrifugation-based method may be applied for obtaining purified sEVs with controlled size, immunoaffinity capturing may be suitable for studies requiring sEVs with high purity but may loss subtypes of sEVs without specific protein marker.

scholarly journals Comparative Evaluation of Methods for Isolating Small Extracellular Vesicles Derived from Pancreatic Cells

2020 ◽  
Author(s):  
Jie-Min Wang ◽  
Yong-Jiang Li ◽  
Jun-Yong Wu ◽  
Jia-Xin Cai ◽  
Jing Wen ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Density Gradient ◽  
Extracellular Vesicles ◽  
Cell Communication ◽  
Size Exclusion ◽  
Density Gradient Ultracentrifugation ◽  
Pancreatic Cancer Cells ◽  
Co Precipitation ◽  
Therapeutic Study

Abstract Background: Small extracellular vesicles (sEVs) are nanosized vesicles involved in cell-to-cell communication. sEVs have been widely studied for clinical applications such as early detection of diseases and as therapeutics. Various methods for sEVs isolation have been using, but different methods may result in different qualities of sEVs and impact downstream analysis and applications. Here, we compared current isolation methods and performed a comparative analysis of sEVs derived from pancreatic cancer cells.Results: Ultracentrifugation, ultrafiltration and co-precipitation as concentration methods were firstly evaluated for yield, size, morphology and protein level of pellets. Then, isolate sEVs obtained by four different purification methods: size exclusion chromatography, density gradient ultracentrifugation, ultracentrifugation, and immunoaffinity capturing, were analysed and compared. For the concentration process, ultracentrifugation method obtained high quality and concentration pellets. For the purification process, immunoaffinity capturing method obtained the purest sEVs with less contaminants, while density gradient ultracentrifugation-based method obtained sEVs with the smallest size. Proteomic analysis revealed distinct protein contents of purified sEVs. Conclusions: For isolating sEVs derived from pancreatic cancer cells, ultracentrifugation-based method is recommended for concentration of sEVs, density gradient ultracentrifugation-based method may be suitable for isolation of sEVs for therapeutic study, immunoaffinity capturing may be applied for studies exploring sEVs as biomarkers.

scholarly journals Cushioned-Density Gradient Ultracentrifugation (C-DGUC) improves the isolation efficiency of extracellular vesicles

PLoS ONE ◽  
2019 ◽  
Vol 14 (4) ◽  
pp. e0215324 ◽  
Author(s):  
Phat Duong ◽  
Allen Chung ◽  
Laura Bouchareychas ◽  
Robert L. Raffai
Keyword(s):  
Density Gradient ◽  
Extracellular Vesicles ◽  
Density Gradient Ultracentrifugation

scholarly journals Comparative evaluation of methods for isolating small extracellular vesicles derived from pancreatic cancer cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Jie-Min Wang ◽  
Yong-Jiang Li ◽  
Jun-Yong Wu ◽  
Jia-Xin Cai ◽  
Jing Wen ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Cancer Cells ◽  
Density Gradient ◽  
Extracellular Vesicles ◽  
Pancreatic Cancer Cell Line ◽  
Size Exclusion ◽  
Density Gradient Ultracentrifugation ◽  
High Concentration ◽  
Protein Marker ◽  
Pancreatic Cancer Cells

Abstract Background Small extracellular vesicles (sEVs) are nanosized vesicles involved in cell-to-cell communication. sEVs have been widely studied for clinical applications such as early detection of diseases and as therapeutics. Various methods for sEVs isolation are been using, but different methods may result in different qualities of sEVs and impact downstream analysis and applications. Here, we compared current isolation methods and performed a comparative analysis of sEVs from supernatant of cultured pancreatic cancer cells. Methods Ultracentrifugation, ultrafiltration and co-precipitation as concentration methods were firstly evaluated for yield, size, morphology and protein level of pellets. Then, isolate sEVs obtained by four different purification methods: size exclusion chromatography, density gradient ultracentrifugation, ultracentrifugation, and immunoaffinity capturing, were analysed and compared. Results For the concentration process, ultracentrifugation method obtained high quality and high concentration of pellets. For the purification process, immunoaffinity capturing method obtained the purest sEVs with less contaminants, while density gradient ultracentrifugation-based method obtained sEVs with the smallest size. Proteomic analysis revealed distinct protein contents of purified sEVs from different methods. Conclusions For isolating sEVs derived from supernatant of cultured pancreatic cancer cell line, ultracentrifugation-based method is recommended for concentration of sEVs, density gradient ultracentrifugation-based method may be applied for obtaining purified sEVs with controlled size, immunoaffinity capturing may be suitable for studies requiring sEVs with high purity but may loss subtypes of sEVs without specific protein marker.

scholarly journals Extracellular Vesicles from Follicular and Ampullary Fluid Isolated by Density Gradient Ultracentrifugation Improve Bovine Embryo Development and Quality

2021 ◽  
Vol 22 (2) ◽  
pp. 578
Author(s):  
Anise Asaadi ◽  
Nima Azari Dolatabad ◽  
Hadi Atashi ◽  
Annelies Raes ◽  
Petra Van Damme ◽  
...  
Keyword(s):  
Density Gradient ◽  
Embryo Development ◽  
Extracellular Vesicles ◽  
Apoptotic Cell ◽  
Developmental Competence ◽  
Size Exclusion ◽  
Density Gradient Ultracentrifugation ◽  
Bovine Embryo ◽  
Step Size

Extracellular vesicles (EVs) have been isolated from follicular (FF) and ampullary oviduct fluid (AOF), using different isolation methods. However, it is not clear whether different purification methods can affect the functionality of resulting EVs. Here, we compared two methods (OptiPrep™ density gradient ultracentrifugation (ODG UC) and single-step size exclusion chromatography (SEC) (qEV IZON™ single column)) for the isolation of EVs from bovine FF and AOF. Additionally, we evaluated whether the addition of EVs derived either by ODG UC or SEC from FF or AOF during oocyte maturation would yield extra benefits for embryo developmental competence. The characterization of EVs isolated using ODG UC or SEC from FF and AOF did not show any differences in terms of EV sizes (40–400 nm) and concentrations (2.4 ± 0.2 × 1012−1.8 ± 0.2 × 1013 particles/mL). Blastocyst yield and quality was higher in groups supplemented with EVs isolated from FF and AOF by ODG UC, with higher total cell numbers and a lower apoptotic cell ratio compared with the other groups (p < 0.05). Supplementing in vitro maturation media with EVs derived by ODG UC from AOF was beneficial for bovine embryo development and quality.

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