Recovery of extracellular vesicles from liquid samples using polyamine solution

Abstract

Liquid biopsies mainly analyze nucleic acids and proteins in the free-state or extracellular vesicles (EVs) in non-solid biological samples, primarily blood. Collecting and processing liquid biopsy samples is challenging due to the large volume of samples and reagents and the need for special equipment. In a previous study, we reported a method for enriching free nucleic acids using a polyamine solution that is effective for liquid biopsy. We also investigated the reactivity of this method for EV recovery in cell culture supernatants using mass spectrometry. Samples were prepared from the cell line NCI-N87 supernatants after 48 h of culture in a serum-free medium. For comparison, samples were treated using a solution containing polyamines (PA method) or ultracentrifugation (UC method). The liquid chromatography-tandem mass spectrometry (LC-MS/MS) using the single-pot solid-phase-enhanced sample-preparation (SP3) method revealed differences between the two methods in the total ion chromatogram of the sample. However, the results of the Gene Ontology (GO) analysis showed that both methods achieved the best enrichment in GO terms related to EV. In addition, the volcano plot analysis revealed that proteins suggested to exist in EVs were distributed in areas consistent with both methods. These results indicated that the PA method can recover EV proteins in liquid samples, and their comprehensive analysis is possible using the SP3 method.