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Size and shape characterization of hydrated and desiccated exosomes


Exosomes are stable nanovesicles secreted by cells into the circulation. Their reported sizes differ substantially, which likely reflects the difference in the isolation techniques used, the cells that secreted them, and the methods used in their characterization. We analyzed the influence of the last factor on the measured sizes and shapes of hydrated and desiccated exosomes isolated from the serum of a pancreatic cancer patient and a healthy control. We found that hydrated exosomes are close-to-spherical nanoparticles with a hydrodynamic radius that is substantially larger than the geometric size. For desiccated exosomes, we found that the desiccated shape and sizing are influenced by the manner in which drying occurred. Isotropic desiccation in aerosol preserves the near-spherical shape of the exosomes, whereas drying on a surface likely distorts their shapes and influences the sizing results obtained by techniques that require surface fixation prior to analysis.

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  1. 1.

    Thery C, Zitvogel L, Amigorena S (2002) Exosomes: composition, biogenesis and function. Nat Rev Immunol 2:569–579

    CAS  Google Scholar 

  2. 2.

    Trams EG, Lauter CJ, Salem N Jr, Heine U (1981) Exfoliation of membrane ecto-enzymes in the form of micro-vesicles. Biochim Biophys Acta 645:63–70

    Article  CAS  Google Scholar 

  3. 3.

    Palma J et al (2012) MicroRNAs are exported from malignant cells in customized particles. Nucleic Acids Res 40:9125–9138

    Article  CAS  Google Scholar 

  4. 4.

    King HW, Michael MZ, Gleadle JM (2012) Hypoxic enhancement of exosome release by breast cancer cells. BMC Cancer 12:421

    Article  CAS  Google Scholar 

  5. 5.

    Kucharzewska P, Belting M (2013) Emerging roles of extracellular vesicles in the adaptive response of tumour cells to microenvironmental stress. J Extracell Vesicles. doi:10.3402/jev.v2i0.20304

    Google Scholar 

  6. 6.

    Chevillet JR et al (2014) Quantitative and stoichiometric analysis of the microRNA content of exosomes. Proc Natl Acad Sci U S A 111:14888–14893

    Article  CAS  Google Scholar 

  7. 7.

    Gallo A, Tandon M, Alevizos I, Illei GG (2012) The majority of microRNAs detectable in serum and saliva is concentrated in exosomes. PLoS ONE 7:e30679

    Article  CAS  Google Scholar 

  8. 8.

    Pucci F, Pittet MJ (2013) Molecular pathways: tumor-derived microvesicles and their interactions with immune cells in vivo. Clin Cancer Res 19:2598–2604

    CAS  Google Scholar 

  9. 9.

    Ge R, Tan E, Sharghi-Namini S, Asada HH (2012) Exosomes in cancer microenvironment and beyond: have we overlooked these extracellular messengers? Cancer Microenviron 5:323–332

    Article  CAS  Google Scholar 

  10. 10.

    Zhang Y et al (2010) Secreted monocytic miR-150 enhances targeted endothelial cell migration. Mol Cell 39:133–144

    Article  CAS  Google Scholar 

  11. 11.

    Regev-Rudzki N et al (2013) Cell-cell communication between malaria-infected red blood cells via exosome-like vesicles. Cell 153:1120–1133

    Article  CAS  Google Scholar 

  12. 12.

    Théry C, Amigorena S, Raposo G, Clayton A (2006) Isolation and characterization of exosomes from cell culture supernatants and biological fluids. Curr Protoc Cell Biol 30:3.22.1–3.22.29

    Google Scholar 

  13. 13.

    Taylor DD, Zacharias W, Gercel-Taylor C (2011) Exosome isolation for proteomic analyses and RNA profiling. Methods Mol Biol 728:235–246

    Article  CAS  Google Scholar 

  14. 14.

    Witwer KW et al (2013) Standardization of sample collection, isolation and analysis methods in extracellular vesicle research. J Extracell Vesicles. doi:10.3402/jev.v2i0.20360

    Google Scholar 

  15. 15.

    Tauro BJ et al (2012) Comparison of ultracentrifugation, density gradient separation, and immunoaffinity capture methods for isolating human colon cancer cell line LIM1863-derived exosomes. Methods 56:293–304

    Article  CAS  Google Scholar 

  16. 16.

    Chen C et al (2010) Microfluidic isolation and transcriptome analysis of serum microvesicles. Lab Chip 10:505–511

    Article  CAS  Google Scholar 

  17. 17.

    Vlassov AV, Magdaleno S, Setterquist R, Conrad R (2012) Exosomes: current knowledge of their composition, biological functions, and diagnostic and therapeutic potentials. Biochim Biophys Acta 1820:940–948

    Article  CAS  Google Scholar 

  18. 18.

    Rekker K et al (2014) Comparison of serum exosome isolation methods for microRNA profiling. Clin Biochem 47:135–138

    Article  CAS  Google Scholar 

  19. 19.

    Alvarez ML, Khosroheidari M, Kanchi Ravi R, DiStefano JK (2012) Comparison of protein, microRNA, and mRNA yields using different methods of urinary exosome isolation for the discovery of kidney disease biomarkers. Kidney Int 82:1024–1032

    Article  CAS  Google Scholar 

  20. 20.

    György B et al (2011) Membrane vesicles, current state-of-the-art: emerging role of extracellular vesicles. Cell Mol Life Sci 68:2667–2688

    Article  Google Scholar 

  21. 21.

    van der Pol E, Coumans F, Varga Z, Krumrey M, Nieuwland R (2013) Innovation in detection of microparticles and exosomes. J Thromb Haemost 11(Suppl 1):36–45

    Google Scholar 

  22. 22.

    van der Pol E et al (2010) Optical and non-optical methods for detection and characterization of microparticles and exosomes. J Thromb Haemost 8:2596–2607

    Article  Google Scholar 

  23. 23.

    Thery C, Ostrowski M, Segura E (2009) Membrane vesicles as conveyors of immune responses. Nat Rev Immunol 9:581–593

    Article  CAS  Google Scholar 

  24. 24.

    Bacher G et al (2001) Charge-reduced nano electrospray ionization combined with differential mobility analysis of peptides, proteins, glycoproteins, noncovalent protein complexes and viruses. J Mass Spectrom 36:1038–1052

    Article  CAS  Google Scholar 

  25. 25.

    Caulfield MP et al (2008) Direct determination of lipoprotein particle sizes and concentrations by ion mobility analysis. Clin Chem 54:1307–1316

    Article  CAS  Google Scholar 

  26. 26.

    Guha S, Pease LF III, Brorson KA, Tarlov MJ, Zachariah MR (2011) Evaluation of electrospray differential mobility analysis for virus particle analysis: potential applications for biomanufacturing. J Virol Methods 178:201–208

    Article  CAS  Google Scholar 

  27. 27.

    Guha S, Li M, Tarlov MJ, Zachariah MR (2012) Electrospray-differential mobility analysis of bionanoparticles. Trends Biotechnol 30:291–300

    Article  CAS  Google Scholar 

  28. 28.

    Pease LF et al (2010) Packing and size determination of colloidal nanoclusters. Langmuir 26:11384–11390

    Article  CAS  Google Scholar 

  29. 29.

    Wiedensohler A (1988) An approximation of the bipolar charge distribution for particles in the submicron size range. J Aerosol Sci 19:387–389

    Article  CAS  Google Scholar 

  30. 30.

    Flagan RC (2008) Differential mobility analysis of aerosols: a tutorial. KONA Powder Part J 26:254–268

    Article  CAS  Google Scholar 

  31. 31.

    Lattin JR, Belnap DM, Pitt WG (2012) Formation of eLiposomes as a drug delivery vehicle. Colloids Surf B 89:93–100

    Article  CAS  Google Scholar 

  32. 32.

    Belnap DM, Grochulski WD, Olson NH, Baker TS (1993) Use of radial density plots to calibrate image magnification for frozen-hydrated specimens. Ultramicroscopy 48:347–358

    Article  CAS  Google Scholar 

  33. 33.

    Deegan RD et al (1997) Capillary flow as the cause of ring stains from dried liquid drops. Nature 389:827–829

    Article  CAS  Google Scholar 

  34. 34.

    Nguyen TAH, Hampton MA, Nguyen AV (2013) Evaporation of nanoparticle droplets on smooth hydrophobic surfaces: the inner coffee ring deposits. J Phys Chem C 117:4707–4716

    Article  CAS  Google Scholar 

  35. 35.

    Yunker PJ, Still T, Lohr MA, Yodh AG (2011) Suppression of the coffee-ring effect by shape-dependent capillary interactions. Nature 476:308–311

    Article  CAS  Google Scholar 

  36. 36.

    Still T, Yunker PJ, Yodh AG (2012) Surfactant-induced Marangoni eddies alter the coffee-rings of evaporating colloidal drops. Langmuir 28:4984–4988

    Article  CAS  Google Scholar 

  37. 37.

    Kaufman SL, Skogen JW, Dorman FD, Zarrin F, Lewis KC (1996) Macromolecule analysis based on electrophoretic mobility in air: globular proteins. Anal Chem 68:1895–1904

    Article  CAS  Google Scholar 

  38. 38.

    György B et al (2011) Detection and isolation of cell-derived microparticles are compromised by protein complexes resulting from shared biophysical parameters. Blood 117:e39–e48

    Article  Google Scholar 

  39. 39.

    Van Deun J et al (2014) The impact of disparate isolation methods for extracellular vesicles on downstream RNA profiling. J Extracell Vesicles. doi:10.3402/jev.v3.24858

    Google Scholar 

  40. 40.

    Caradec J et al (2014) Reproducibility and efficiency of serum-derived exosome extraction methods. Clin Biochem 47:1286–1292

    Article  CAS  Google Scholar 

  41. 41.

    Jeppesen DK et al (2014) Comparative analysis of discrete exosome fractions obtained by differential centrifugation. J Extracell Vesicles. doi:10.3402/jev.v3.25011

    Google Scholar 

  42. 42.

    Mathivanan S, Simpson RJ (2009) ExoCarta: a compendium of exosomal proteins and RNA. Proteomics 9:4997–5000

    Article  CAS  Google Scholar 

  43. 43.

    Laulagnier K et al (2005) Characterization of exosome subpopulations from RBL-2H3 cells using fluorescent lipids. Blood Cells Mol Dis 35:116–121

    Article  CAS  Google Scholar 

  44. 44.

    Bobrie A, Colombo M, Krumeich S, Raposo G, Théry C (2012) Diverse subpopulations of vesicles secreted by different intracellular mechanisms are present in exosome preparations obtained by differential ultracentrifugation. J Extracell Vesicles. doi:10.3402/jev.v1i0.18397

    Google Scholar 

  45. 45.

    Yellon DM, Davidson SM (2014) Exosomes: nanoparticles involved in cardioprotection? Circ Res 114:325–332

    Article  CAS  Google Scholar 

  46. 46.

    Kobayashi M et al (2014) Ovarian cancer cell invasiveness is associated with discordant exosomal sequestration of Let-7 miRNA and miR-200. J Transl Med 12:4

    Article  Google Scholar 

  47. 47.

    Petersen KE et al (2014) A review of exosome separation techniques and characterization of B16-F10 mouse melanoma exosomes with AF4-UV-MALS-DLS-TEM. Anal Bioanal Chem 406:7855–7866

    Article  CAS  Google Scholar 

  48. 48.

    Sharma S, Gillespie BM, Palanisamy V, Gimzewski JK (2011) Quantitative nanostructural and single-molecule force spectroscopy biomolecular analysis of human-saliva-derived exosomes. Langmuir 27:14394–14400

    Article  CAS  Google Scholar 

  49. 49.

    Sharma S et al (2010) Structural-mechanical characterization of nanoparticle exosomes in human saliva, using correlative AFM, FESEM, and force spectroscopy. ACS Nano 4:1921–1926

    Article  CAS  Google Scholar 

  50. 50.

    Sharma S, Das K, Woo J, Gimzewski JK (2014) Nanofilaments on glioblastoma exosomes revealed by peak force microscopy. J R Soc Interface 11:20131150

    Article  Google Scholar 

  51. 51.

    Conde-Vancells J et al (2008) Characterization and comprehensive proteome profiling of exosomes secreted by hepatocytes. J Proteome Res 7:5157–5166

    Article  CAS  Google Scholar 

  52. 52.

    Zhou Y et al (2013) Exosomes released by human umbilical cord mesenchymal stem cells protect against cisplatin-induced renal oxidative stress and apoptosis in vivo and in vitro. Stem Cell Res Ther 4:34

    Article  CAS  Google Scholar 

  53. 53.

    Coleman BM, Hanssen E, Lawson VA, Hill AF (2012) Prion-infected cells regulate the release of exosomes with distinct ultrastructural features. FASEB J 26:4160–4173

    Article  CAS  Google Scholar 

  54. 54.

    van der Pol E, Böing AN, Harrison P, Sturk A, Nieuwland R (2012) Classification, functions, and clinical relevance of extracellular vesicles. Pharmacol Rev 64:676–705

    Article  Google Scholar 

  55. 55.

    Momen-Heravi F et al (2012) Impact of biofluid viscosity on size and sedimentation efficiency of the isolated microvesicles. Front Physiol 3:162

    CAS  Google Scholar 

  56. 56.

    Sokolova V et al (2011) Characterisation of exosomes derived from human cells by nanoparticle tracking analysis and scanning electron microscopy. Colloids Surf B 87:146–150

    Article  CAS  Google Scholar 

  57. 57.

    Tscharnuter WT (2006) Photon correlation spectroscopy in particle sizing. In Encyclopedia of analytical chemistry. Wiley, Hoboken. doi:10.1002/9780470027318.a1512

    Google Scholar 

  58. 58.

    Besseling NAM (1997) Theory of hydration forces between surfaces. Langmuir 13:2113–2122

    Article  CAS  Google Scholar 

  59. 59.

    He L, Hu Y, Wang M, Yin Y (2012) Determination of solvation layer thickness by a magnetophotonic approach. ACS Nano 6:4196–4202

    Article  CAS  Google Scholar 

  60. 60.

    Tathireddy P, Choi Y-H, Skliar M (2008) Particle AC electrokinetics in planar interdigitated microelectrode geometry. J Electrostat 66:609–619

    Article  CAS  Google Scholar 

  61. 61.

    Iyer S, Gaikwad RM, Subba-Rao V, Woodworth CD, Sokolov I (2009) Atomic force microscopy detects differences in the surface brush of normal and cancerous cells. Nat Nanotechnol 4:389–393

    Article  CAS  Google Scholar 

  62. 62.

    Frank J (2002) Single-particle imaging of macromolecules by cryo-electron microscopy. Annu Rev Biophys Biomol Struct 31:303–319

    Article  CAS  Google Scholar 

  63. 63.

    Oh E et al (2011) Cellular uptake and fate of PEGylated gold nanoparticles is dependent on both cell-penetration peptides and particle size. ACS Nano 5:6434–6448

    Article  CAS  Google Scholar 

  64. 64.

    Lavialle F et al (2009) Nanovesicles released by Dictyostelium cells: a potential carrier for drug delivery. Int J Pharm 380:206–215

    Article  CAS  Google Scholar 

  65. 65.

    Varga Z et al (2014) Towards traceable size determination of extracellular vesicles. J Extracell Vesicles. doi:10.3402/jev.v3.23298

    Google Scholar 

  66. 66.

    Hardij J et al (2013) Characterisation of tissue factor-bearing extracellular vesicles with AFM: comparison of air-tapping-mode AFM and liquid peak force AFM. J Extracell Vesicles. doi:10.3402/jev.v2i0.21045

    Google Scholar 

  67. 67.

    Dragovic RA et al (2011) Sizing and phenotyping of cellular vesicles using nanoparticle tracking analysis. Nanomedicine 7:780–788

    Article  CAS  Google Scholar 

  68. 68.

    Momen-Heravi F et al (2012) Alternative methods for characterization of extracellular vesicles. Front Physiol 3:354

    Google Scholar 

  69. 69.

    Hood JL, Pan H, Lanza GM, Wickline SA (2009) Paracrine induction of endothelium by tumor exosomes. Lab Investig 89:1317–1328

    Article  Google Scholar 

  70. 70.

    Atay S, Gercel-Taylor C, Kesimer M, Taylor DD (2011) Morphologic and proteomic characterization of exosomes released by cultured extravillous trophoblast cells. Exp Cell Res 317:1192–1202

    Article  CAS  Google Scholar 

  71. 71.

    Tian T, Wang Y, Wang H, Zhu Z, Xiao Z (2010) Visualizing of the cellular uptake and intracellular trafficking of exosomes by live-cell microscopy. J Cell Biochem 111:488–496

    Article  CAS  Google Scholar 

  72. 72.

    Tatischeff I, Larquet E, Falcón-Pérez JM, Turpin P-Y, Kruglik SG (2012) Fast characterisation of cell-derived extracellular vesicles by nanoparticles tracking analysis, cryo-electron microscopy, and Raman tweezers microspectroscopy. J Extracell Vesicles. doi:10.3402/jev.v1i0.19179

    Google Scholar 

  73. 73.

    Wahlgren J, Karlson TDL, Glader P, Telemo, Valadi H (2012) Activated human T cells secrete exosomes that participate in IL-2 mediated immune response signaling. PLoS ONE 7:e49723

    Article  CAS  Google Scholar 

  74. 74.

    Ng YH et al (2013) Endometrial exosomes/microvesicles in the uterine microenvironment: a new paradigm for embryo-endometrial cross talk at implantation. PLoS ONE 8:e58502

    Article  CAS  Google Scholar 

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The authors are indebted to Brian F. Woodfield of Brigham Young University (Department of Chemistry) for providing access to the NTA instrument. The authors acknowledge financial support from the National Science Foundation (award number IGERT-0903715) and the University of Utah (Department of Chemical Engineering Seed Grant and the Graduate Research Fellowship award).

Author contributions

M.S. and P.S.B. conceived the study, R.R. isolated exosomes, V.S.C. and R.R. performed NTA measurements, DLS measurements were performed by V.S.C. and M.S, D.M.B performed cryo-TEM imaging, SEM imaging was performed by Y.J. and V.S.C., electrospray DMA measurements were performed by Y.H.T., V.S.C., L.F.P., and M.S., V.S.C. and M.S. analyzed the experimental results; A.E.B. wrote the MATLAB code to analyze the imaged exosomes, M.S. performed statistical analysis, and V.S.C. and K.J.B. performed manual sizing of the imaged exosomes. The manuscript was written by V.S.C. and M.S., and was edited by all authors.

Conflict of interest

The authors declare that they have no competing financial interests.

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Correspondence to Mikhail Skliar.

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Chernyshev, V.S., Rachamadugu, R., Tseng, Y.H. et al. Size and shape characterization of hydrated and desiccated exosomes. Anal Bioanal Chem 407, 3285–3301 (2015).

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  • Hydrated and desiccated exosomes
  • Size and shape characterization