Measurement of refractive index by nanoparticle tracking analysis reveals heterogeneity in extracellular vesicles

Chris Gardiner; Michael Shaw; Patrick Hole; Jonathan Smith; Dionne Tannetta; Christopher W. Redman; Ian L. Sargent

doi:10.3402/jev.v3.25361

Journal of Extracellular Vesicles (Nov 2014)

Measurement of refractive index by nanoparticle tracking analysis reveals heterogeneity in extracellular vesicles

Chris Gardiner,
Michael Shaw,
Patrick Hole,
Jonathan Smith,
Dionne Tannetta,
Christopher W. Redman,
Ian L. Sargent

Affiliations

Chris Gardiner: Nuffield Department of Obstetrics and Gynaecology, University of Oxford, Level 3 Women's Centre, John Radcliffe Hospital, Oxford, UK
Michael Shaw: Nuffield Department of Obstetrics and Gynaecology, University of Oxford, Level 3 Women's Centre, John Radcliffe Hospital, Oxford, UK
Patrick Hole: Malvern Instruments, Minton Park, Amesbury, UK
Jonathan Smith: Malvern Instruments, Minton Park, Amesbury, UK
Dionne Tannetta: Nuffield Department of Obstetrics and Gynaecology, University of Oxford, Level 3 Women's Centre, John Radcliffe Hospital, Oxford, UK
Christopher W. Redman: Nuffield Department of Obstetrics and Gynaecology, University of Oxford, Level 3 Women's Centre, John Radcliffe Hospital, Oxford, UK
Ian L. Sargent: Nuffield Department of Obstetrics and Gynaecology, University of Oxford, Level 3 Women's Centre, John Radcliffe Hospital, Oxford, UK

DOI: https://doi.org/10.3402/jev.v3.25361
Journal volume & issue: Vol. 3, no. 0
pp. 1 – 6

Abstract

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Introduction: Optical techniques are routinely used to size and count extracellular vesicles (EV). For comparison of data from different methods and laboratories, suitable calibrators are essential. A suitable calibrator must have a refractive index (RI) as close to that of EV as possible but the RI of EV is currently unknown. To measure EV, RI requires accurate knowledge of size and light scattering. These are difficult to measure as most EVs cannot be resolved by light microscopy and their diameter is smaller than the wavelength of visible light. However, nanoparticle tracking analysis (NTA) provides both size and relative light scattering intensity (rLSI) values. We therefore sought to determine whether it was possible to use NTA to measure the RI of individual EVs. Methods: NTA was used to measure the rLSI and size of polystyrene and silica microspheres of known size and RI (1.470 and 1.633, respectively) and of EV isolated from a wide range of cells. We developed software, based on Mie scattering code, to calculate particle RI from the rLSI data. This modelled theoretical scattering intensities for polystyrene and silica microspheres of known size (100 and 200 nm) and RI. The model was verified using data from the polystyrene and silica microspheres. Size and rLSI data for each vesicle were processed by the software to generate RI values. Results: The following modal RI measurements were obtained: fresh urinary EV 1.374, lyophilised urinary EV 1.367, neuroblastoma EV 1.393, blood EV 1.398, EV from activated platelets 1.390, small placental EV 1.364–1.375 and 1.398–1.414 for large placental EV (>200 nm). Large placental EV had a significantly higher RI than small placental EV (p1.40 were observed for some large (>200 nm) microvesicles. Conclusion: This method for measuring EV RI will be useful for developing appropriate calibrators for EV measurement.

Published in Journal of Extracellular Vesicles

ISSN: 2001-3078 (Online)
Publisher: Wiley
Country of publisher: United States
LCC subjects: Science: Biology (General): Cytology
Website: https://onlinelibrary.wiley.com/journal/20013078

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