title: Multispectral Optical Tweezers for Biochemical Fingerprinting of CD9-Positive Exosome Subpopulations.
abstract: Extracellular vesicles (EVs), including exosomes, are circulating nanoscale particles heavily implicated in cell signaling and can be isolated in vast numbers from human biofluids. Study of their molecular profiling and materials properties is currently underway for purposes of describing a variety of biological functions and diseases. However, the large, and as yet largely unquantified, variety of EV subpopulations differing in composition, size, and likely function necessitates characterization schemes capable of measuring single vesicles. Here we describe the first application of multispectral optical tweezers (MS-OTs) to single vesicles for molecular fingerprinting of EV subpopulations. This versatile imaging platform allows for sensitive measurement of Raman chemical composition (e.g., variation in protein, lipid, cholesterol, nucleic acids), coupled with discrimination by fluorescence markers. For exosomes isolated by ultracentrifugation, we use MS-OTs to interrogate the CD9-positive subpopulations via antibody fluorescence labeling and Raman spectra measurement. We report that the CD9-positive exosome subset exhibits reduced component concentration per vesicle and reduced chemical heterogeneity compared to the total purified EV population. We observed that specific vesicle subpopulations are present across exosomes isolated from cell culture supernatant of several clonal varieties of mesenchymal stromal cells and also from plasma and ascites isolated from human ovarian cancer patients.
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California Davis , Sacramento, California 95817, United States.
authors.forename: Randy P
authors.identifier: 0000-0001-8193-1664
authors.initials: RP
authors.lastname: Carney
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California Davis , Sacramento, California 95817, United States.
authors.forename: Sidhartha
authors.initials: S
authors.lastname: Hazari
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California Davis , Sacramento, California 95817, United States.
authors.forename: Macalistair
authors.initials: M
authors.lastname: Colquhoun
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California Davis , Sacramento, California 95817, United States.
authors.forename: Di
authors.initials: D
authors.lastname: Tran
authors.affiliation: Department of Surgery, University of Washington , 850 Republican Street, Seattle, Washington 98195-0005, United States.
authors.forename: Billanna
authors.initials: B
authors.lastname: Hwang
authors.affiliation: Department of Surgery, University of Washington , 850 Republican Street, Seattle, Washington 98195-0005, United States.
authors.forename: Michael S
authors.initials: MS
authors.lastname: Mulligan
authors.affiliation: Department of Bioengineering, University of Washington , 3720 15th Avenue NE, Seattle, Washington 98195, United States.
authors.forename: James D
authors.initials: JD
authors.lastname: Bryers
authors.affiliation: Division of Gynecologic Oncology, University of California Davis Medical Center , Sacramento, California 98517, United States.
authors.forename: Eugenia
authors.initials: E
authors.lastname: Girda
authors.affiliation: Division of Gynecologic Oncology, University of California Davis Medical Center , Sacramento, California 98517, United States.
authors.forename: Gary S
authors.initials: GS
authors.lastname: Leiserowitz
authors.affiliation: Department of Precision Machinery and Precision Instrumentation, University of Science and Technology of China , Hefei, Anhui 230027 China.
authors.forename: Zachary J
authors.initials: ZJ
authors.lastname: Smith
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California Davis , Sacramento, California 95817, United States.|Division of Hematology/Oncology, University of California Davis Cancer Center , Sacramento, California 95817, United States.
authors.forename: Kit S
authors.initials: KS
authors.lastname: Lam
mesh_terms: D000818:Animals; D000906:Antibodies / Q000276:immunology; D055354:Exosomes / Q000378:metabolism*; D005260:Female; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000378:metabolism; D053758:Nanoparticles / Q000737:chemistry; D052898:Optical Tweezers; D010051:Ovarian Neoplasms / Q000378:metabolism / Q000473:pathology; D025341:Principal Component Analysis; D051381:Rats; D013059:Spectrum Analysis, Raman; D060245:Tetraspanin 29 / Q000032:analysis* / Q000276:immunology
chemical_list: D000906:Antibodies; D005456:Fluorescent Dyes; D060245:Tetraspanin 29