Sensitive Separation Device Enables Diagnostic Analysis of Cancer Exosomes

Exosomes are cell-derived vesicles that are present in many and perhaps all biological fluids, including blood, urine, and cultured medium of cell cultures. The reported diameter of exosomes is between 30 and 100 nanometers, which is larger than low-density lipoproteins but much smaller than red blood cells. Exosomes, which contain RNA, proteins, lipids, and metabolites that are reflective of the cell type of origin, are either released from the cell when multivesicular bodies (MVBs) fuse with the plasma membrane, or they are released directly from the plasma membrane. Exosomes have specialized functions and play a key role in coagulation, intercellular signaling, and waste management.

Investigators at Washington State University (Pullman, USA) sought to overcome the technical barriers hindering the diagnostic applications of cancer exosomes by developing a paper-based isotachophoresis (ITP) technology capable of 1) rapid isolation and identification of exosomes from both malignant and healthy cells and 2) multiplex detection of selected exosomal protein biomarkers of the target exosomes.

While generally not achieving the resolution of other forms of electrophoresis, ITP has been successfully employed for difficult samples, such as very small peptides, not amenable to traditional techniques. ITP has also shown great promise for the analysis of complex mixtures of molecules of different classes. Although, technically, isotachophoresis separates samples by electrophoretic mobility, the layers of sample molecules move at the same speed.

The novel technology described in the current study combined the focusing power of ITP with the multiplex capability of paper-based lateral flow to achieve on-board separation of target exosomes from large extracellular vesicles, followed by electrokinetic enrichment of the targets. This created an ultrasensitive platform for comprehensive exosome analysis.

For a proof of concept, the technology platform was tested with human serum samples spiked with exosomes derived from healthy human serum and a prostate cancer cell line. Results revealed that under anionic ITP conditions, the device showed superior performance in simultaneous detection of the cancer exosomes and normal exosomes with an observed limit of detection more than 30-fold better than that of enhanced ELISA. In a subsequent step, the technology was shown to be capable of the rapid profiling of a selected protein biomarker panel associated with the target exosomes.

"This has the potential to become a technique capable of concentrating samples by orders of magnitude in minutes," said senior author Dr Wenji Dong, associate professor of chemical engineering and bioengineering at Washington State University. "Exosomes provide a unique opportunity as a cancer marker."

The exosome concentration device was described in the May 15, 2020, online edition of the journal Biosensors and Bioelectronics.

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