The samples were centrifuged at 300× g for 10 min, followed by centrifugation at 3000× g for 10 min. The clarified supernatant was then concentrated to approximately 500 μL on a 100 KD Amicon Ultra Centrifugal filter (Millipore, Temecula, CA, USA). The exosomes were then isolated from the concentrated supernatant by size exclusion chromatography (SEC). Sephacryl S-300 High Resolution (GE Healthcare, Chicago, IL, USA) was packed on a glass econo-column chromatography column (BioRad, Hercules, CA, USA) (10 cm height, 1.5 cm diameter). The column was washed with 0.32% Sodium Citrate in PBS, and the supernatant was loaded onto the column and allowed to enter the resin by gravity flow. The eluate was collected in 23 fractions of 15 drops (~500 μL) on a Model 2110 Fraction Collector (BioRad, Hercules, CA, USA). For each fraction, the presence of exosomes was determined by nanoparticle tracking analysis as described below. The exosome-containing fractions were then further concentrated to 1/100th of the original supernatant.
Glass econo column chromatography column
Manufactured by Bio-Rad
Sourced in United States
The Glass econo-column chromatography column is a laboratory equipment used for separation and purification of various biomolecules, such as proteins, nucleic acids, and small molecules, through the process of column chromatography. It is a glass column designed to facilitate the flow of mobile phase through a stationary phase, allowing the components of a mixture to be separated based on their differences in interaction with the stationary phase.
Automatically generated - may contain errors
3 protocols using glass econo column chromatography column
1
Exosome Isolation by Size Exclusion Chromatography
2
Isolation of NK Cell Exosomes
3
Synthesis and Purification of Butyrate-Modified Fructooligosaccharides
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B-FOS was produced by Bifido Inc. (Gangwon-do, Korea). It was synthesized by adding butyrate (Sigma-Aldrich, St. Louis, MO, USA) to an aqueous solution of >90% pure fructooligosaccharides (FOS, Samyang Corp., Seoul, Korea). FOS and butyric anhydride 98% (Sigma-Aldrich, St. Louis, MO, USA) were used in the B-FOS synthesis process. FOS (50% w/v) was mixed with butyrate in a ratio of 10: 1, at a temperature of 50 °C for 3 h. The product was neutralized with a NaOH solution to complete the reaction. The purification of B-FOS was performed by filtration through synthetic absorbent Diaion HP20 packed in a 50 × 5 cm Glass Econo-Column chromatography column (Bio-Rad, Hercules, CA, USA). The 40%–70% (v/v) ethanol eluent was collected and the purity of B-FOS was tested by thin-layer chromatography (TLC) with a solvent of 1-propanol/water/ethyl acetate (7:2:1, v/v) (See Figure S1 ). Purified B-FOS was concentrated via a speed vacuum concentrator (ScanSpeed 40, Labogene, Lynge, Denmark) and freeze-dried (Ilshin Biobase, Yangju, Korea). The final B-FOS product was a soluble carbohydrate powder.
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