Pre-analytical Methods Optimization for Extracellular Vesicle Isolation and Measurement
Extracellular RNA (exRNA) has been widely studied for use as a biomarker for early detection and disease monitoring. ExRNA is released by cells into biofluids such as plasma, urine, saliva, and cerebrospinal fluid (CSF). Plasma is a common biofluid collected and used for research and clinical studies due to its minimally invasive nature. The current method for plasma collection post venipuncture is to perform one centrifugal spin to separate the plasma from other blood components which includes white blood cells and platelets prior to freeze banking. Our aim is to reduce cellular contaminants through two successive centrifugal spins prior to banking in an effort to increase RNA diversity. Using RNA sequencing we observed a decrease in ribosomal RNA (rRNA), more lncRNA, coding genes, and a higher variety of mappable genes from the two successive spin plasma samples compared to the traditional single spin. This supports that an additional spin prior to plasma banking will reduce cellular contamination, a main source of rRNA, and increase the diversity of detectable RNAs. We would recommend, based on our data, that plasma samples destined for exRNA experiments, should include a double spin before freezing and banking.