Journal: Current Issues in Molecular Biology

Article Title: Methodological and Short-Term Diurnal Variation in Surface and Cargo Proteins in Plasma Extracellular Vesicles

doi: 10.3390/cimb48010120

Figure Lengend Snippet: Morphologic and protein characterization of EV samples. ( A ) TEM and IEM images of isolated EV preparations. TEM showing vesicular structures around 200 nm. IEM images showing CD9+ extracellular vesicles with marker antibodies connected to the outer membrane with some debris present. Yellow bar: 200 nm. All images acquired at 20,000×, Panels displayed at the same magnification. Yellow arrows indicate objects of interest. ( B ) Western blot of pooled participant samples confirming the presence of CD9 and ALIX and lipoprotein marker Apo-B.

Article Snippet: To facilitate the phenotyping of EVs, a total of 17 antibodies were selected: CD9 (Ancell, Bayport, MN, USA, Cat.# 156-020, Clone: SN4/C3-3A2), CD4 (R&D Systems, Minneapolis, MN, USA, Cat.# MAB379, Clone: 34930), CD63 (Biorad, Hercules, CA, USA, Cat.# MCA2142, Clone: MEM-259), CD8a (R&D Systems, Cat.# MAB1509, Clone: 37006), CD81 (Ancell, Cat.# 302-020, Clone: 1.3.3.22), HLA ABC (Biolegend, San Diego, CA, USA, Cat.# 311402, Clone: W6/32), CD142 (R&D Systems, Cat#. MAB2339, Clone: 323514), Flotillin-1 (Abcam, Cambridge, UK, Cat.# ab41927) CD42a (LS Bio, Seattle, WA, USA, Cat.# LS- C45240 ), CD45 (R&D Systems Cat.# MAB1430, Clone: 2D1), CD106 (R&D Systems Cat.# MAB809, Clone:HAE-2Z), MIC A/B (R&D Systems Cat.# MAB13001 Clone: 159207), ICAM-1 (eBioscience, San Diego, CA, USA, Cat.# BMS1011, Clone: R6.5), HLA DR/DP/DQ (Capricobio, Norcross, GA, USA, Cat.# 104701 Clone: HB-145/IVA12), CD31 (R&D Systems, Cat.# AF806, Polyclonal, TNF RII (R&D Systems, Cat.# MAB726 Clone: 22210) and TNF RI (R&D Systems, Cat.# MAB225, Clone:16803).

Techniques: Isolation, Marker, Membrane, Western Blot