Questioning the purity of the media - Extracellular small non-coding RNA contaminants in fetal bovine serum and serum-free media

In the research field of extracellular vesicles (EVs), the use of EV-depleted fetal bovine serum (FBS) for in vitro studies is highly recommended to eliminate the confounding effects of media derived EVs. EV-depleted FBS may either be prepared by ultracentrifugation or bought commercially, nevertheless these depletion methods do not guarantee an RNA-free preparation. In this study we have addressed the RNA contamination issue in FBS, ultracentrifuged EV-depleted FBS, commercially available EV-depleted FBS, and also from our recently developed filtration based EV depleted FBS. Commercially available serum-free, xeno-free defined media were also screened for RNA contamination. Small non-coding RNA profiles of extracellular vesicles derived from different types of cell media were generated by next-generation sequencing. Samples included FBS (n=2), commercial exosome-depleted FBS (n=2), ultracentrifuged EV-depleted FBS (n=2), ultrafiltrated EV-depleted FBS (n=2) and defined serum-free medium (n=1)