Physically Isolated OptiPrep (TM) Fractions for exRNA Analysis

An integrative analysis of human biofluid data in the exRNA Atlas revealed the existence of distinct extracellular RNA cargo types. In order to detect differences in density between cargo types, cushioned density gradient ultracentrifugation (C-DGUC) of serum and plasma was performed using OptiPrem (TM) density gradient. 250 ml whole blood was collected from five adult female and five adult male consenting donors supervised by Dr. Louise C. Laurent at the University of California, San Diego, Department of Obstetrics, Gynecology, and Reproductive Sciences and Sanford Consortium for Regenerative Medicine. These samples were utilized to generate the serum and plasma OptiPrep (TM) fractions (1-3, 4-7, 9-12) and whole biofluid. Plasma and serum were fractionated using cushioned density gradient ultracentrifugation (C-DGUC) via OptiPrep (TM) density gradient. RNA was then isolated and sequenced, and western blot and mass spectrometry analyses were performed. 78 of 80 samples were used in the exRNA data analysis (2 samples were not used due to low read count) RNA-Seq raw data is to be made available through dbGaP (controlled access) due to patient privacy concerns.