Introduction: Current methods of isolating extracelluar vesicles such as ultracentrifugation can be time consuming, requiring specialised equipment, labour intensive, and quality control can be difficult to achieve. Therefore, the goal of this study is to develop a highly-reproducible, automated method to isolate extracellular vesicles using high-performance liquid chromatography (HPLC) without human intervention.
Method: A modular HPLC system with a fraction collector coupled with a multimodal chromatography column was used for this study. Each purification round consists of three cycles, 1) injection of sample, 2) removal of bound contaminating proteins, 3) equilibration of column. To optimise the HPLC method, multiple replicates of fetal bovine serum followed by human serum were processed, with multiple elution fractions collected from each starting sample. The successful isolation of extracellular vesicles were confirmed as per the International Society for Extracellular Vesicles guidelines.
Results: A highly reproducible method of isolating extracellular vesicles from both bovine and human serum was achieved. In addition, from one biological sample, using the optimised automated method, the majority of the total vesicles can be purified into only one tube. Furthermore, the purity of the isolated vesicles was comparable to current isolation techniques.
Conclusions: The use of automated systems to isolate extracellular vesicles could have important implications in the field of diagnostics where high-throughput and reproducibility is essential.