Nanoscale imaging of RNA with expansion microscopy

Fei Chen, Asmamaw T. Wassie, Allison J. Cote, Anubhav Sinha, Shahar Alon, Shoh Asano, Evan R. Daugharthy, Jae Byum Chang, Adam Marblestone, George M. Church, Arjun Raj, Edward S. Boyden

Research output: Contribution to journalArticlepeer-review

273 Scopus citations

Abstract

The ability to image RNA identity and location with nanoscale precision in intact tissues is of great interest for defining cell types and states in normal and pathological biological settings. Here, we present a strategy for expansion microscopy of RNA. We developed a small-molecule linker that enables RNA to be covalently attached to a swellable polyelectrolyte gel synthesized throughout a biological specimen. Then, postexpansion, fluorescent in situ hybridization (FISH) imaging of RNA can be performed with high yield and specificity as well as single-molecule precision in both cultured cells and intact brain tissue. Expansion FISH (ExFISH) separates RNAs and supports amplification of single-molecule signals (i.e., via hybridization chain reaction) as well as multiplexed RNA FISH readout. ExFISH thus enables super-resolution imaging of RNA structure and location with diffraction-limited microscopes in thick specimens, such as intact brain tissue and other tissues of importance to biology and medicine.

Original languageEnglish
Pages (from-to)679-684
Number of pages6
JournalNature Methods
Volume13
Issue number8
DOIs
StatePublished - Aug 2016
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2016 Nature America, Inc.

Funding

Lightsheet imaging was performed in the W.M. Keck Facility for Biological Imaging at the Whitehead Institute for Biomedical Research. We would like to acknowledge W. Salmon for assistance with the Zeiss Z.1 lightsheet, S. Olenych from Carl Zeiss Microscopy for providing the microscopy filters, and H.T. Choi and N. Pierce for advice and consultation on HCR. E.R.D. is supported by NIH CEGS grant P50 HG005550, NIH CEGS grant 1 RM1 HG008525, and NSF GRF grant DGE1144152. A.T.W. acknowledges the Hertz Foundation Fellowship. F.C. acknowledges the NSF Fellowship and Poitras Fellowship. AR and AC acknowledge support from NIH/NHLBI grant 1U01HL129998. E.S.B. acknowledges support by the New York Stem Cell Foundation'Robertson Award, NSF CBET 1053233, MIT Media Lab Consortium, the MIT Synthetic Intelligence Project, NIH Director's Pioneer Award 1DP1NS087724, NIH 2R01DA029639, NIH Director's Transformative Award 1R01MH103910, NIH 1R24MH106075, IARPA D16PC00008, the Open Philanthropy Project, and Jeremy and Joyce Wertheimer. J.-B.C. was supported by a Simons Postdoctoral Fellowship.

FundersFunder number
MIT Media Lab Consortium
MIT Synthetic Intelligence Project1R24MH106075, 1R01MH103910, 1DP1NS087724, 2R01DA029639
Whitehead Institute for Biomedical Research
National Science FoundationDGE1144152
National Institutes of Health1 RM1 HG008525, R24MH106075, P50HG005550, R01MH103910, R01DA029639
National Heart, Lung, and Blood InstituteU01HL129998
Simons Foundation
New York Stem Cell FoundationCBET 1053233
Hertz Foundation
Intelligence Advanced Research Projects ActivityD16PC00008

    Fingerprint

    Dive into the research topics of 'Nanoscale imaging of RNA with expansion microscopy'. Together they form a unique fingerprint.

    Cite this