Non-reversible tissue fixation retains extracellular vesicles for in situ imaging

Mrinali P. Gupta; Sangeetha Tandalam; Shariss Ostrager; Alexander S. Lever; Angus R. Fung; David D. Hurley; Gemstonn B. Alegre; Jasmin E. Espinal; H. Lawrence Remmel; Sushmita Mukherjee; Benjamin M. Levine; Russell P. Robins; Henrik Molina; Brian D. Dill; Candia M. Kenific; Thomas Tuschl; David Lyden; Donald J. D’Amico; John T.G. Pena

doi:10.1038/s41592-019-0623-4

Non-reversible tissue fixation retains extracellular vesicles for in situ imaging

Mrinali P. Gupta, Sangeetha Tandalam, Shariss Ostrager, Alexander S. Lever, Angus R. Fung, David D. Hurley, Gemstonn B. Alegre, Jasmin E. Espinal, H. Lawrence Remmel, Sushmita Mukherjee, Benjamin M. Levine, Russell P. Robins, Henrik Molina, Brian D. Dill, Candia M. Kenific, Thomas Tuschl, David Lyden, Donald J. D’Amico, John T.G. Pena^*

^*Corresponding author for this work

Department of Pathology

Research output: Contribution to journal › Article › Academic › peer-review

Abstract

Extracellular vesicles (EVs) are secreted nanosized particles with many biological functions and pathological associations. The inability to image EVs in fixed tissues has been a major limitation to understanding their role in healthy and diseased tissue microenvironments. Here, we show that crosslinking mammalian tissues with formaldehyde results in significant EV loss, which can be prevented by additional fixation with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for visualization of EVs in a range of normal and cancer tissues.

Original language	English
Pages (from-to)	1269-1273
Number of pages	5
Journal	Nature Methods
Volume	16
Issue number	12
DOIs	https://doi.org/10.1038/s41592-019-0623-4
Publication status	Published - 1 Dec 2019

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Gupta, M. P., Tandalam, S., Ostrager, S., Lever, A. S., Fung, A. R., Hurley, D. D., Alegre, G. B., Espinal, J. E., Remmel, H. L., Mukherjee, S., Levine, B. M., Robins, R. P., Molina, H., Dill, B. D., Kenific, C. M., Tuschl, T., Lyden, D., D’Amico, D. J., & Pena, J. T. G. (2019). Non-reversible tissue fixation retains extracellular vesicles for in situ imaging. Nature Methods, 16(12), 1269-1273. https://doi.org/10.1038/s41592-019-0623-4

@article{2ed15db3d40248fab4165879a86bc0ab,

title = "Non-reversible tissue fixation retains extracellular vesicles for in situ imaging",

abstract = "Extracellular vesicles (EVs) are secreted nanosized particles with many biological functions and pathological associations. The inability to image EVs in fixed tissues has been a major limitation to understanding their role in healthy and diseased tissue microenvironments. Here, we show that crosslinking mammalian tissues with formaldehyde results in significant EV loss, which can be prevented by additional fixation with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for visualization of EVs in a range of normal and cancer tissues.",

author = "Gupta, {Mrinali P.} and Sangeetha Tandalam and Shariss Ostrager and Lever, {Alexander S.} and Fung, {Angus R.} and Hurley, {David D.} and Alegre, {Gemstonn B.} and Espinal, {Jasmin E.} and Remmel, {H. Lawrence} and Sushmita Mukherjee and Levine, {Benjamin M.} and Robins, {Russell P.} and Henrik Molina and Dill, {Brian D.} and Kenific, {Candia M.} and Thomas Tuschl and David Lyden and D{\textquoteright}Amico, {Donald J.} and Pena, {John T.G.}",

note = "Funding Information: We thank J. Moore for his technical support for this manuscript. This work was performed in part at the Weill Cornell Medicine Imaging Core (for Electron Microscopy), and Genomics Resources Core, Cornell NanoScale Facility, a member of the National Nanotechnology Coordinated Infrastructure (NNCI), which is supported by the National Science Foundation (Grant NNCI-1542081). The Heed Foundation supported M.G. and J.T.G.P. The Rockefeller University Proteomics Resource Center acknowledges funding from the Leona M. and Harry B. Helmsley Charitable Trust for mass spectrometer instrumentation. T.T. was supported by an Extracellular RNA Communication grants U19CA179564 from the NIH. D.L. was supported by NCI R35 CA232093 Outstanding Investigator Award from the NIH. J.T.G.P. supported by Daedalus Fund at Weill Cornell Medicine, The Shulsky Foundation, Research to Prevent Blindness Foundation and Knights Templar Eye Foundation Inc. Publisher Copyright: {\textcopyright} 2019, The Author(s), under exclusive licence to Springer Nature America, Inc.",

year = "2019",

month = dec,

day = "1",

doi = "10.1038/s41592-019-0623-4",

language = "English",

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pages = "1269--1273",

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Gupta, MP, Tandalam, S, Ostrager, S, Lever, AS, Fung, AR, Hurley, DD, Alegre, GB, Espinal, JE, Remmel, HL, Mukherjee, S, Levine, BM, Robins, RP, Molina, H, Dill, BD, Kenific, CM, Tuschl, T, Lyden, D, D’Amico, DJ & Pena, JTG 2019, 'Non-reversible tissue fixation retains extracellular vesicles for in situ imaging', Nature Methods, vol. 16, no. 12, pp. 1269-1273. https://doi.org/10.1038/s41592-019-0623-4

TY - JOUR

T1 - Non-reversible tissue fixation retains extracellular vesicles for in situ imaging

AU - Gupta, Mrinali P.

AU - Tandalam, Sangeetha

AU - Ostrager, Shariss

AU - Lever, Alexander S.

AU - Fung, Angus R.

AU - Hurley, David D.

AU - Alegre, Gemstonn B.

AU - Espinal, Jasmin E.

AU - Remmel, H. Lawrence

AU - Mukherjee, Sushmita

AU - Levine, Benjamin M.

AU - Robins, Russell P.

AU - Molina, Henrik

AU - Dill, Brian D.

AU - Kenific, Candia M.

AU - Tuschl, Thomas

AU - Lyden, David

AU - D’Amico, Donald J.

AU - Pena, John T.G.

N1 - Funding Information: We thank J. Moore for his technical support for this manuscript. This work was performed in part at the Weill Cornell Medicine Imaging Core (for Electron Microscopy), and Genomics Resources Core, Cornell NanoScale Facility, a member of the National Nanotechnology Coordinated Infrastructure (NNCI), which is supported by the National Science Foundation (Grant NNCI-1542081). The Heed Foundation supported M.G. and J.T.G.P. The Rockefeller University Proteomics Resource Center acknowledges funding from the Leona M. and Harry B. Helmsley Charitable Trust for mass spectrometer instrumentation. T.T. was supported by an Extracellular RNA Communication grants U19CA179564 from the NIH. D.L. was supported by NCI R35 CA232093 Outstanding Investigator Award from the NIH. J.T.G.P. supported by Daedalus Fund at Weill Cornell Medicine, The Shulsky Foundation, Research to Prevent Blindness Foundation and Knights Templar Eye Foundation Inc. Publisher Copyright: © 2019, The Author(s), under exclusive licence to Springer Nature America, Inc.

PY - 2019/12/1

Y1 - 2019/12/1

N2 - Extracellular vesicles (EVs) are secreted nanosized particles with many biological functions and pathological associations. The inability to image EVs in fixed tissues has been a major limitation to understanding their role in healthy and diseased tissue microenvironments. Here, we show that crosslinking mammalian tissues with formaldehyde results in significant EV loss, which can be prevented by additional fixation with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for visualization of EVs in a range of normal and cancer tissues.

AB - Extracellular vesicles (EVs) are secreted nanosized particles with many biological functions and pathological associations. The inability to image EVs in fixed tissues has been a major limitation to understanding their role in healthy and diseased tissue microenvironments. Here, we show that crosslinking mammalian tissues with formaldehyde results in significant EV loss, which can be prevented by additional fixation with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) for visualization of EVs in a range of normal and cancer tissues.

UR - http://www.scopus.com/inward/record.url?scp=85075138621&partnerID=8YFLogxK

U2 - 10.1038/s41592-019-0623-4

DO - 10.1038/s41592-019-0623-4

M3 - Article

C2 - 31712780

AN - SCOPUS:85075138621

SN - 1548-7091

VL - 16

SP - 1269

EP - 1273

JO - Nature Methods

JF - Nature Methods

IS - 12

ER -

Non-reversible tissue fixation retains extracellular vesicles for in situ imaging

Abstract

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