title: Comparative Analysis of Technologies for Quantifying Extracellular Vesicles (EVs) in Clinical Cerebrospinal Fluids (CSF).
abstract: Extracellular vesicles (EVs) have emerged as a promising biomarker platform for glioblastoma patients. However, the optimal method for quantitative assessment of EVs in clinical bio-fluid remains a point of contention. Multiple high-resolution platforms for quantitative EV analysis have emerged, including methods grounded in diffraction measurement of Brownian motion (NTA), tunable resistive pulse sensing (TRPS), vesicle flow cytometry (VFC), and transmission electron microscopy (TEM). Here we compared quantitative EV assessment using cerebrospinal fluids derived from glioblastoma patients using these methods. For EVs <150 nm in diameter, NTA detected more EVs than TRPS in three of the four samples tested. VFC particle counts are consistently 2-3 fold lower than NTA and TRPS, suggesting contribution of protein aggregates or other non-lipid particles to particle count by these platforms. While TEM yield meaningful data in terms of the morphology, its particle count are consistently two orders of magnitude lower relative to counts generated by NTA and TRPS. For larger particles (>150 nm in diameter), NTA consistently detected lower number of EVs relative to TRPS. These results unveil the strength and pitfalls of each quantitative method alone for assessing EVs derived from clinical cerebrospinal fluids and suggest that thoughtful synthesis of multi-platform quantitation will be required to guide meaningful clinical investigations.
authors.affiliation: Center for Theoretical and Applied Neuro-Oncology, University of California San Diego, San Diego, California, United States of America.
authors.forename: Johnny C
authors.initials: JC
authors.lastname: Akers
authors.affiliation: Center for Theoretical and Applied Neuro-Oncology, University of California San Diego, San Diego, California, United States of America.
authors.forename: Valya
authors.initials: V
authors.lastname: Ramakrishnan
authors.affiliation: Scintillon Institute for Biomedical and Bioenergy Research, San Diego, California, United States of America.
authors.forename: John P
authors.initials: JP
authors.lastname: Nolan
authors.affiliation: Scintillon Institute for Biomedical and Bioenergy Research, San Diego, California, United States of America.
authors.forename: Erika
authors.initials: E
authors.lastname: Duggan
authors.affiliation: Izon Science, Christchurch, New Zealand.
authors.forename: Chia-Chun
authors.initials: CC
authors.lastname: Fu
authors.affiliation: Neurology Service, Massachusetts General Hospital, and Program in Neuroscience, Harvard Medical School, Boston, Massachusetts, United States of America.
authors.forename: Fred H
authors.initials: FH
authors.lastname: Hochberg
authors.affiliation: Center for Theoretical and Applied Neuro-Oncology, University of California San Diego, San Diego, California, United States of America.
authors.forename: Clark C
authors.initials: CC
authors.lastname: Chen
authors.affiliation: Center for Theoretical and Applied Neuro-Oncology, University of California San Diego, San Diego, California, United States of America.
authors.forename: Bob S
authors.initials: BS
authors.lastname: Carter
mesh_terms: D002555:Cerebrospinal Fluid / Q000378:metabolism*; D019411:Clinical Laboratory Techniques / Q000592:standards*; D000067128:Extracellular Vesicles; D005434:Flow Cytometry; D006801:Humans; D066298:In Vitro Techniques; D046529:Microscopy, Electron, Transmission