SmartSEC-DeLipo Single EV Isolation System

Product Description

Elevate your small extracellular vesicle (sEV) isolation game with our cutting-edge SmartSEC-DeLipo^TM kit, engineered to provide unmatched purity and quality while efficiently depleting lipoprotein contaminants. Harnessing the power of SmartSEC^TM coupled with innovative lipoprotein removal technology, this kit streamlines the sEV isolation process, ensuring pure sEVs samples for downstream analyses.

• New generation of mixed-mode chromatography
• Advanced lipoprotein depletion
• Efficient workflow
• High yield and purity
• Versatile compatibility with various applications
• Superior performance

The SmartSEC-DeLipo Kit utilizes SmartSEC^TM columns, a new generation of mixed-mode chromatography, which provides the dual functionality of size exclusion and affinity interaction modes. Combining the two modes within a single resin media eliminates the time consuming and labor-intensive processes of classical SEC, delivering highly purified sEV samples. The kit is compatible with various sample types containing lipoprotein including plasma, serum, breast milk et al, ensuring versatility and flexibility in experimental design.

The SmartSEC-DeLipo Kit is also available in a 96-well HT format. Standard versions of the kit and separate DeLipo lipoprotein removal columns are listed below under Related Products.

Importance of Removing Lipoproteins from sEV

Extracellular vesicles (EVs) from blood are of great importance to understand the biological role of circulating EVs and to develop EVs as biomarkers of disease. However, isolating EVs from blood poses challenges due to the simultaneous presence of lipoprotein particles. Lipoproteins may interfere with downstream analyses or functional assays performed on EVs. For instance, lipoproteins contain microRNAs as their cargos. In studies investigating the biological roles of EVs or their potential as biomarkers, the presence of lipoproteins could confound the results or mask the true effects of EVs. Recently, several studies showed that ultracentrifugation or size exclusion chromatography (SEC) alone cannot completely separate blood EVs from lipoprotein particles. Therefore, improved separation of EVs from lipoproteins is crucial for a detailed functional analysis of circulating EVs, thus making blood a viable source for EV biomarker discovery.

How it works

The SmartSEC-DeLipo workflow is fast and easy. Simply apply 250 µL of cleared serum or plasma with additional column buffer or up to 0.5 mL of other biofluids directly to the pre-washed column, incubate, and centrifuge to elute the Evs then go through DeLipo column to remove lipoprotein from the isolated sEVs.

Supporting Data

Figure 2. sEV isolated with SmartSEC-DeLipo is much purer. (A) sEV isolated from 250ul of serum with SmartSEC-DeLipo shows higher expression of EV markers, such as CD9, CD81 and TSG101 than those isolated with SmartSEC single when same amount of EV protein were applied for western blot. When same volume of eluted sEV samples were applied for western blot, EV markers’ expression is about the same for sEV isolated from 250ul of serum with SmartSEC-DeLipo or SmartSEC single. (B) sEV isolated from 250ul of plasma or breast milk with SmartSEC-DeLipo shows higher expression of EV markers, such as CD9, CD81 and TSG101 than those isolated with SmartSEC single when same amount of EV proteins was applied for western blot. (C) fNTA data shows that sEV particle number were decreased in plasma or breast milk sample when isolated with SmartSEC-DeLipo in comparison with that isolated with SmartSEC single. However, when purity was evaluated with EV particle number per same amount of EV protein, sEV isolated with SmartSEC-DeLipo demonstrates higher purity than those isolated with SmartSEC single.

Figure 3. Most of the Lipoproteins are removed in sEV isolated with SmartSEC-DeLipo. (A) ApoB and ApoE lipoproteins’ concentrations were evaluated in serum sample, sEV isolated from serum with SmartSEC single and sEV isolated from serum with SmartSEC-DeLipo by ELISA. SmarSECTM single dramatically reduced Lipoproteins in isolated sEV and SmartSEC-DeLipo further deplete the remaining Lipoproteins in yield sEV. (B) Percentage of ApoB and ApoE depletion was calculated based on ApoB and ApoE concentration in sEV isolated with SmartSEC-DeLipo vs. SmartSEC single from serum, plasma and breast milk samples.

Figure 4. sEV isolated with SmartSEC-DeLipo shows typical EV morphology. Transmission electron microscopy (TEM) of sEVs isolated from serum using SmartSEC-DeLipo possess typical EV morphology—intact vesicles with a double layer of membranes.

Figure 5. sEV isolated with SmartSEC-DeLipo is biologically functional to delivery small RNA cargo to the recipient cells. (A) Cy3-labeled control siRNA were loaded without or with sEV isolated from serum with SmartSEC single or SmartSEC-DeLipo using Exo-Fect siRNA/miRNA reagent (EXFT200A-1) and delivered to HEK 293 cells. Cells imaged 36-48 hours post transfection. Shown are the bright-field images (bottom row), and fluorescence images (top row). Both sEV isolated with SmartSEC single or SmartSEC-DeLipo can efficiently transfer Cy3-labeled siRNA to most of the imaged cells. (B) HPRT siRNA were loaded without or with sEV isolated from serum with SmartSEC single or SmartSEC-DeLipo using Exo-Fect siRNA/miRNA reagent (EXFT200A-1) and delivered to HEK 293 cells. Cells were lysed and applied for western blot 48 hours post transfection. Both sEV isolated with SmartSEC single or SmartSEC-DeLipo can efficiently transfer HPRT siRNA to HEK 293 cells and achieve knockdown of the endogenous HPRT expression.