Western Blot Analysis of HMDM Exosomes

Western blot was performed as described previously [87 (link)]. Lysates of HMDMs or their EVs were prepared in RIPA buffer supplemented with cOmplete Protease Inhibitor Cocktail (Roche, Basel, Switzerland). Anti-Alix (1A12, Santa Cruz Biotechnology, Dallas, Texas, USA), anti-CD63 (Santa Cruz Biotechnology), anti-Calnexin (AF18, Santa Cruz Biotechnology), anti-cytochrome c (A-8, Santa Cruz Biotechnology), and anti-TLR7 (4F4, sc-57463, Santa Cruz Biotechnology) were used as primary antibodies.

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Pawar K., Shigematsu M., Sharbati S, & Kirino Y. (2020). Infection-induced 5′-half molecules of tRNAHisGUG activate Toll-like receptor 7. PLoS Biology, 18(12), e3000982.

Publication 2020

cOmplete Protease Inhibitor Cocktail anti-CD63 anti-Calnexin (AF18, anti-cytochrome c (A-8,

Antibodies Buffer Calnexin Cytochrome c Protease inhibitor Ripa Western blot

Corresponding Organization : Freie Universität Berlin

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Variable analysis

independent variables

Lysates of HMDMs or their EVs

dependent variables

Protein expression levels of Alix, CD63, Calnexin, cytochrome c, and TLR7

control variables

RIPA buffer supplemented with cOmplete Protease Inhibitor Cocktail (Roche, Basel, Switzerland)

controls

Positive controls: None specified
Negative controls: None specified

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