Figure 1. Typical Western Blot Analyses of EVs Isolated from Human Plasma and Mesenchymal Stromal Cell (MSC)-Conditioned Medium

The Western blot analyses in the above examples show positive isolation of EVs with CD9, CD63, and CD81 tetraspanin markers from (A) human plasma and (B) MSC-conditioned medium. Western blots were run under non-reducing conditions.

Figure 2. Tetraspanin Protein Expression and Recovery of EVs Using EasySep™ Human Pan/CD9/CD63/CD81 Extracellular Vesicle Positive Selection Kits

(A) EVs were isolated from plasma using either EasySep™ Human Extracellular Vesicle Positive Selection Kits or differential ultracentrifugation (2 x 70 min, 100,000 xg) and isolated fractions were analyzed by western blot for tetraspanin protein expression. (B) Equal or higher recovery of EVs was achieved from mesenchymal stromal cell (MSC)-conditioned MesenCult™-ACF Plus Medium and plasma using EasySep™ Human PanExtracellular Vesicle Positive Selection Kit when compared to EVs isolated using other commercially available immunocapture-based EV isolation kits.

Figure 3. Images of Plasma-Derived EVs Isolated Using EasySep™ Human Pan-Extracellular Vesicle Positive Selection Kit

Transmission electron microscopy (TEM) analysis following EasySep™ positive selection shows intact and spherical-shaped (arrows) plasma-derived EVs. The isolated EVs are attached to tetrameric antibody complexes and magnetic particles.

Figure 4. Top 10 Cellular Compartment Gene Ontology Terms for EVs Isolated from Plasma or MSC-Conditioned Medium Using EasySep™ Human Pan-Extracellular Vesicle Positive Selection Kit

Proteins present in EVs isolated from (A) plasma and (B) MSC-conditioned medium using EasySep™ Human Pan-Extracellular Vesicle Positive Selection Kit were detected by proteomic analysis. \Gene Ontology analysis showed the detected proteins were grouped by terms associated with EVs, confirming the quality and compatibility of isolated EVs with mass spectrometry analysis.

Figure 5. Common microRNAs (miRNAs) in Plasma-Derived EVs Isolated Using EasySep™ Human Pan-Extracellular Vesicle Positive Selection Kit

MicroRNAs (miRNAs) found in plasma were detected with RT-qPCR demonstrating the compatibility of EasySep™ Human Pan-Extracellular Vesicle Positive Selection Kit with downstream RNA extraction and RNA analysis. The increase in Ct value following EV lysis using 0.1% Triton™X-100 and RNase digestion demonstrate the integrity of isolated EVs.