Abstract
Extracellular Vesicles (EVs) are membrane-derived vesicles continuously shed by cells into the extracellular space. With cargos containing nucleic acids and other proteins, EVs play key roles in intercellular communication and other biological processes and are widely regarded as promising biomarkers for disease detection. EVs are abundantly found in virtually every biofluid, including blood, urine, and saliva. While there are numerous technologies to isolate EVs in blood and urine, isolating EVs from saliva remains a challenge, as it is highly complex and heterogeneous—cells, debris, and other proteins can all inhibit the isolation of EVs by traditional platforms. Here, we employ the CandyCollect, a lollipop-inspired sampling device with open microfluidic channels, as a non-invasive and patient-friendly alternative for the capture of salivary EVs. The CandyCollect simplifies sample preparation by effectively pre-concentrating EVs on the device surface before EVs are eluted off of the CandyCollect, labeled with cholesterol-tagged oligonucleotides, and subsequently detected by qPCR with primers specific for the tagged oligos to enumerate the relative number of EVs. We demonstrate that downstream EV cargo analysis can be performed using Simoa. Overall, the CandyCollect ushers a new method to capture, enumerate, and analyze salivary EVs.
Competing Interest Statement
Ashleigh B. Theberge, Xiaojing Su, Erwin Berthier, and Sanitta Thongpang filed patent 63/152,103 (International Publication Number: WO 2022/178291 Al) through the University of Washington on the CandyCollect oral sampling device. Erwin Berthier and Ashleigh B. Theberge filed patent 63/683,571 through the University of Washington on a related platform. Ashleigh B. Theberge reports filing multiple patents through the University of Washington and receiving a gift to support research outside the submitted work from Ionis Pharmaceuticals. Erwin Berthier is an inventor on multiple patents filed by Tasso, Inc., the University of Washington, and the University of Wisconsin. Sanitta Thongpang has ownership in Salus Discovery, LLC, and Tasso, Inc. Erwin Berthier has ownership in Salus Discovery, LLC, and Tasso, Inc. and is employed by Tasso, Inc. However, this research is not related to these companies. Sanitta Thongpang, Erwin Berthier, and Ashleigh B. Theberge have ownership in Seabright, LLC, which will advance new tools for diagnostics and clinical research, potentially including the CandyCollect device. The terms of this arrangement have been reviewed and approved by the University of Washington in accordance with its policies governing outside work and financial conflicts of interest in research. Lydia L. Sohn has filed a patent 62/942,216 through the University of California, Berkeley on the DNA labeling of extracellular vesicles for enumeration. The other authors have no conflicts of interest to disclose.
Footnotes
Updated author list; Section on Simoa updated to clarify methods used
