Figure 5.

Extracellular vesicles (EV) Long Interspersed Element-1 (LINE-1) levels in individual human plasma donors. LINE-1 mRNA and ORF1p levels were quantified in ostensibly healthy donors using plasma volumes that could be commonly acquired by venipuncture. For each donor, EVs were isolated from 2.5 mL plasma via ultracentrifugation. Thirty micrograms of EV protein from six different donors (D1–D6) was used for Western blotting. (A) Ponceau staining of the membrane demonstrates equal protein loading. Whole plasma from Donor 1 was included as a reference. (B) Immunoblot of EV proteins and ORF1p. (C) RNA was extracted from EVs and then subjected to One-Step RT-qPCR. Mean, SEM shown. To examine LINE-1 mRNA expression more closely, mRNA levels from all donors used in this study were converted to log2 FC values (D,E) and a one-sample t-test was used to determine if levels were significantly greater than zero (* p < 0.05).