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. 2020 May 18;23(6):101182. doi: 10.1016/j.isci.2020.101182

Figure 3.

Figure 3

Isolation of Total RNA from EV- and No-EV-Enriched Plasma Fractions for RNA Sequencing

(A) Total RNA from extracellular vesicles (EVs) isolated from either pooled or individual plasma samples was used to compare gene expression between no-EV (fractions 1–5) and EV-enriched (fractions 6–10) plasma compartments. The different sizes of EVs were isolated by cushioned-density gradient ultracentrifugation using iodixanol and purified with dialysis prior to RNA extraction. RNA-sequencing libraries were constructed in duplicates using the SMARTer Pico v2 kit and subsequently sequenced on Illumina's HiSeq 2500 platform.

(B) Western blot analysis of CD9, CD63, and Alix markers for extracellular vesicles, AGO2 for Argonaute 2 proteins, and APOA1 for high-density lipoproteins in all fractions (1–12) retrieved from iodixanol cushioned-density gradient ultracentrifugation and dialysis of plasma samples.