---
_id: '7941'
abstract:
- lang: eng
text: Expansion microscopy is a recently developed super-resolution imaging technique,
which provides an alternative to optics-based methods such as deterministic approaches
(e.g. STED) or stochastic approaches (e.g. PALM/STORM). The idea behind expansion
microscopy is to embed the biological sample in a swellable gel, and then to expand
it isotropically, thereby increasing the distance between the fluorophores. This
approach breaks the diffraction barrier by simply separating the emission point-spread-functions
of the fluorophores. The resolution attainable in expansion microscopy is thus
directly dependent on the separation that can be achieved, i.e. on the expansion
factor. The original implementation of the technique achieved an expansion factor
of fourfold, for a resolution of 70–80 nm. The subsequently developed X10 method
achieves an expansion factor of 10-fold, for a resolution of 25–30 nm. This technique
can be implemented with minimal technical requirements on any standard fluorescence
microscope, and is more easily applied for multi-color imaging than either deterministic
or stochastic super-resolution approaches. This renders X10 expansion microscopy
a highly promising tool for new biological discoveries, as discussed here, and
as demonstrated by several recent applications.
article_processing_charge: No
author:
- first_name: Sven M
full_name: Truckenbrodt, Sven M
id: 45812BD4-F248-11E8-B48F-1D18A9856A87
last_name: Truckenbrodt
- first_name: Silvio O.
full_name: Rizzoli, Silvio O.
last_name: Rizzoli
citation:
ama: 'Truckenbrodt SM, Rizzoli SO. Simple multi-color super-resolution by X10 microscopy.
In: Methods in Cell Biology. Vol 161. Elsevier; 2021:33-56. doi:10.1016/bs.mcb.2020.04.016'
apa: Truckenbrodt, S. M., & Rizzoli, S. O. (2021). Simple multi-color super-resolution
by X10 microscopy. In Methods in Cell Biology (Vol. 161, pp. 33–56). Elsevier.
https://doi.org/10.1016/bs.mcb.2020.04.016
chicago: Truckenbrodt, Sven M, and Silvio O. Rizzoli. “Simple Multi-Color Super-Resolution
by X10 Microscopy.” In Methods in Cell Biology, 161:33–56. Elsevier, 2021.
https://doi.org/10.1016/bs.mcb.2020.04.016.
ieee: S. M. Truckenbrodt and S. O. Rizzoli, “Simple multi-color super-resolution
by X10 microscopy,” in Methods in Cell Biology, vol. 161, Elsevier, 2021,
pp. 33–56.
ista: 'Truckenbrodt SM, Rizzoli SO. 2021.Simple multi-color super-resolution by
X10 microscopy. In: Methods in Cell Biology. vol. 161, 33–56.'
mla: Truckenbrodt, Sven M., and Silvio O. Rizzoli. “Simple Multi-Color Super-Resolution
by X10 Microscopy.” Methods in Cell Biology, vol. 161, Elsevier, 2021,
pp. 33–56, doi:10.1016/bs.mcb.2020.04.016.
short: S.M. Truckenbrodt, S.O. Rizzoli, in:, Methods in Cell Biology, Elsevier,
2021, pp. 33–56.
date_created: 2020-06-07T22:00:55Z
date_published: 2021-01-01T00:00:00Z
date_updated: 2021-03-11T08:49:08Z
day: '01'
department:
- _id: JoDa
doi: 10.1016/bs.mcb.2020.04.016
external_id:
pmid:
- '33478696'
intvolume: ' 161'
language:
- iso: eng
month: '01'
oa_version: None
page: 33-56
pmid: 1
publication: Methods in Cell Biology
publication_identifier:
isbn:
- 978012820807-6
issn:
- 0091-679X
publication_status: published
publisher: Elsevier
quality_controlled: '1'
scopus_import: '1'
status: public
title: Simple multi-color super-resolution by X10 microscopy
type: book_chapter
user_id: 2DF688A6-F248-11E8-B48F-1D18A9856A87
volume: 161
year: '2021'
...
---
_id: '6052'
abstract:
- lang: eng
text: 'Expansion microscopy is a relatively new approach to super-resolution imaging
that uses expandable hydrogels to isotropically increase the physical distance
between fluorophores in biological samples such as cell cultures or tissue slices.
The classic gel recipe results in an expansion factor of ~4×, with a resolution
of 60–80 nm. We have recently developed X10 microscopy, which uses a gel that
achieves an expansion factor of ~10×, with a resolution of ~25 nm. Here, we provide
a step-by-step protocol for X10 expansion microscopy. A typical experiment consists
of seven sequential stages: (i) immunostaining, (ii) anchoring, (iii) polymerization,
(iv) homogenization, (v) expansion, (vi) imaging, and (vii) validation. The protocol
presented here includes recommendations for optimization, pitfalls and their solutions,
and detailed guidelines that should increase reproducibility. Although our protocol
focuses on X10 expansion microscopy, we detail which of these suggestions are
also applicable to classic fourfold expansion microscopy. We exemplify our protocol
using primary hippocampal neurons from rats, but our approach can be used with
other primary cells or cultured cell lines of interest. This protocol will enable
any researcher with basic experience in immunostainings and access to an epifluorescence
microscope to perform super-resolution microscopy with X10. The procedure takes
3 d and requires ~5 h of actively handling the sample for labeling and expansion,
and another ~3 h for imaging and analysis.'
article_processing_charge: No
article_type: original
author:
- first_name: Sven M
full_name: Truckenbrodt, Sven M
id: 45812BD4-F248-11E8-B48F-1D18A9856A87
last_name: Truckenbrodt
- first_name: Christoph M
full_name: Sommer, Christoph M
id: 4DF26D8C-F248-11E8-B48F-1D18A9856A87
last_name: Sommer
orcid: 0000-0003-1216-9105
- first_name: Silvio O
full_name: Rizzoli, Silvio O
last_name: Rizzoli
- first_name: Johann G
full_name: Danzl, Johann G
id: 42EFD3B6-F248-11E8-B48F-1D18A9856A87
last_name: Danzl
orcid: 0000-0001-8559-3973
citation:
ama: Truckenbrodt SM, Sommer CM, Rizzoli SO, Danzl JG. A practical guide to optimization
in X10 expansion microscopy. Nature Protocols. 2019;14(3):832–863. doi:10.1038/s41596-018-0117-3
apa: Truckenbrodt, S. M., Sommer, C. M., Rizzoli, S. O., & Danzl, J. G. (2019).
A practical guide to optimization in X10 expansion microscopy. Nature Protocols.
Nature Publishing Group. https://doi.org/10.1038/s41596-018-0117-3
chicago: Truckenbrodt, Sven M, Christoph M Sommer, Silvio O Rizzoli, and Johann
G Danzl. “A Practical Guide to Optimization in X10 Expansion Microscopy.” Nature
Protocols. Nature Publishing Group, 2019. https://doi.org/10.1038/s41596-018-0117-3.
ieee: S. M. Truckenbrodt, C. M. Sommer, S. O. Rizzoli, and J. G. Danzl, “A practical
guide to optimization in X10 expansion microscopy,” Nature Protocols, vol.
14, no. 3. Nature Publishing Group, pp. 832–863, 2019.
ista: Truckenbrodt SM, Sommer CM, Rizzoli SO, Danzl JG. 2019. A practical guide
to optimization in X10 expansion microscopy. Nature Protocols. 14(3), 832–863.
mla: Truckenbrodt, Sven M., et al. “A Practical Guide to Optimization in X10 Expansion
Microscopy.” Nature Protocols, vol. 14, no. 3, Nature Publishing Group,
2019, pp. 832–863, doi:10.1038/s41596-018-0117-3.
short: S.M. Truckenbrodt, C.M. Sommer, S.O. Rizzoli, J.G. Danzl, Nature Protocols
14 (2019) 832–863.
date_created: 2019-02-24T22:59:20Z
date_published: 2019-03-01T00:00:00Z
date_updated: 2023-08-24T14:48:33Z
day: '01'
ddc:
- '570'
department:
- _id: JoDa
- _id: Bio
doi: 10.1038/s41596-018-0117-3
ec_funded: 1
external_id:
isi:
- '000459890700008'
pmid:
- '30778205'
file:
- access_level: open_access
checksum: 7efb9951e7ddf3e3dcc2fb92b859c623
content_type: application/vnd.openxmlformats-officedocument.wordprocessingml.document
creator: kschuh
date_created: 2021-06-29T14:41:46Z
date_updated: 2021-06-29T14:41:46Z
file_id: '9619'
file_name: 181031_Truckenbrodt_ExM_NatProtoc.docx
file_size: 84478958
relation: main_file
success: 1
file_date_updated: 2021-06-29T14:41:46Z
has_accepted_license: '1'
intvolume: ' 14'
isi: 1
issue: '3'
language:
- iso: eng
month: '03'
oa: 1
oa_version: Submitted Version
page: 832–863
pmid: 1
project:
- _id: 260C2330-B435-11E9-9278-68D0E5697425
call_identifier: H2020
grant_number: '754411'
name: ISTplus - Postdoctoral Fellowships
- _id: 265CB4D0-B435-11E9-9278-68D0E5697425
call_identifier: FWF
grant_number: I03600
name: Optical control of synaptic function via adhesion molecules
publication: Nature Protocols
publication_status: published
publisher: Nature Publishing Group
quality_controlled: '1'
scopus_import: '1'
status: public
title: A practical guide to optimization in X10 expansion microscopy
type: journal_article
user_id: 4359f0d1-fa6c-11eb-b949-802e58b17ae8
volume: 14
year: '2019'
...
---
_id: '145'
abstract:
- lang: eng
text: Aged proteins can become hazardous to cellular function, by accumulating molecular
damage. This implies that cells should preferentially rely on newly produced ones.
We tested this hypothesis in cultured hippocampal neurons, focusing on synaptic
transmission. We found that newly synthesized vesicle proteins were incorporated
in the actively recycling pool of vesicles responsible for all neurotransmitter
release during physiological activity. We observed this for the calcium sensor
Synaptotagmin 1, for the neurotransmitter transporter VGAT, and for the fusion
protein VAMP2 (Synaptobrevin 2). Metabolic labeling of proteins and visualization
by secondary ion mass spectrometry enabled us to query the entire protein makeup
of the actively recycling vesicles, which we found to be younger than that of
non-recycling vesicles. The young vesicle proteins remained in use for up to ~
24 h, during which they participated in recycling a few hundred times. They were
afterward reluctant to release and were degraded after an additional ~ 24–48 h.
We suggest that the recycling pool of synaptic vesicles relies on newly synthesized
proteins, while the inactive reserve pool contains older proteins.
acknowledgement: We thank Reinhard Jahn for providing a plasmid for YFP-SNAP25. We
thank Erwin Neher for help with the development of the mathematical model of the
synaptic vesicle life cycle. We thank Martin Meschkat, Andreas Höbartner, Annedore
Punge, and Peer Hoopmann for help with the experiments. We thank Burkhard Rammner
for providing the illustrations of synaptic vesicle and protein dynamics. We thank
Manuel Maidorn, Martin Helm, and Katharina N. Richter for critically reading the
manuscript. S.T. was supported by an Excellence Stipend of the Göttingen Graduate
School for Neurosciences, Biophysics, and Molecular Biosciences (GGNB). E.F.F. is
a recipient of long-term fellowships from the European Molecular Biology Organization
(ALTF_797-2012) and from the Human Frontier Science Program (HFSP_LT000830/2013).
The work was supported by grants to S.O.R. from the European Research Council (ERC-2013-CoG
NeuroMolAnatomy) and from the Deutsche Forschungsgemeinschaft (Cluster of Excellence
Nanoscale Microscopy and Molecular Physiology of the Brain, SFB1190/P09, SFB889/A05,
and SFB1286/A03, and DFG RI 1967 7/1). The nanoSIMS instrument was funded by the
German Federal Ministry of Education and Research (03F0626A).
article_number: e98044
article_processing_charge: No
article_type: original
author:
- first_name: Sven M
full_name: Truckenbrodt, Sven M
id: 45812BD4-F248-11E8-B48F-1D18A9856A87
last_name: Truckenbrodt
- first_name: Abhiyan
full_name: Viplav, Abhiyan
last_name: Viplav
- first_name: Sebsatian
full_name: Jähne, Sebsatian
last_name: Jähne
- first_name: Angela
full_name: Vogts, Angela
last_name: Vogts
- first_name: Annette
full_name: Denker, Annette
last_name: Denker
- first_name: Hanna
full_name: Wildhagen, Hanna
last_name: Wildhagen
- first_name: Eugenio
full_name: Fornasiero, Eugenio
last_name: Fornasiero
- first_name: Silvio
full_name: Rizzoli, Silvio
last_name: Rizzoli
citation:
ama: Truckenbrodt SM, Viplav A, Jähne S, et al. Newly produced synaptic vesicle
proteins are preferentially used in synaptic transmission. The EMBO Journal.
2018;37(15). doi:10.15252/embj.201798044
apa: Truckenbrodt, S. M., Viplav, A., Jähne, S., Vogts, A., Denker, A., Wildhagen,
H., … Rizzoli, S. (2018). Newly produced synaptic vesicle proteins are preferentially
used in synaptic transmission. The EMBO Journal. Wiley. https://doi.org/10.15252/embj.201798044
chicago: Truckenbrodt, Sven M, Abhiyan Viplav, Sebsatian Jähne, Angela Vogts, Annette
Denker, Hanna Wildhagen, Eugenio Fornasiero, and Silvio Rizzoli. “Newly Produced
Synaptic Vesicle Proteins Are Preferentially Used in Synaptic Transmission.” The
EMBO Journal. Wiley, 2018. https://doi.org/10.15252/embj.201798044.
ieee: S. M. Truckenbrodt et al., “Newly produced synaptic vesicle proteins
are preferentially used in synaptic transmission,” The EMBO Journal, vol.
37, no. 15. Wiley, 2018.
ista: Truckenbrodt SM, Viplav A, Jähne S, Vogts A, Denker A, Wildhagen H, Fornasiero
E, Rizzoli S. 2018. Newly produced synaptic vesicle proteins are preferentially
used in synaptic transmission. The EMBO Journal. 37(15), e98044.
mla: Truckenbrodt, Sven M., et al. “Newly Produced Synaptic Vesicle Proteins Are
Preferentially Used in Synaptic Transmission.” The EMBO Journal, vol. 37,
no. 15, e98044, Wiley, 2018, doi:10.15252/embj.201798044.
short: S.M. Truckenbrodt, A. Viplav, S. Jähne, A. Vogts, A. Denker, H. Wildhagen,
E. Fornasiero, S. Rizzoli, The EMBO Journal 37 (2018).
date_created: 2018-12-11T11:44:52Z
date_published: 2018-08-01T00:00:00Z
date_updated: 2023-09-13T09:02:48Z
day: '01'
ddc:
- '570'
department:
- _id: JoDa
doi: 10.15252/embj.201798044
external_id:
isi:
- '000440416900005'
pmid:
- '29950309'
file:
- access_level: open_access
checksum: a540feb6c9af6aefc78de531461a8835
content_type: application/pdf
creator: dernst
date_created: 2018-12-17T14:17:29Z
date_updated: 2020-07-14T12:44:56Z
file_id: '5710'
file_name: 2018_EMBO_Truckenbrodt.pdf
file_size: 2846470
relation: main_file
file_date_updated: 2020-07-14T12:44:56Z
has_accepted_license: '1'
intvolume: ' 37'
isi: 1
issue: '15'
language:
- iso: eng
license: https://creativecommons.org/licenses/by/4.0/
month: '08'
oa: 1
oa_version: Published Version
pmid: 1
publication: The EMBO Journal
publication_identifier:
issn:
- 0261-4189
publication_status: published
publisher: Wiley
publist_id: '7778'
quality_controlled: '1'
scopus_import: '1'
status: public
title: Newly produced synaptic vesicle proteins are preferentially used in synaptic
transmission
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1
volume: 37
year: '2018'
...
---
_id: '6499'
abstract:
- lang: eng
text: Expansion microscopy is a recently introduced imaging technique that achieves
super‐resolution through physically expanding the specimen by ~4×, after embedding
into a swellable gel. The resolution attained is, correspondingly, approximately
fourfold better than the diffraction limit, or ~70 nm. This is a major improvement
over conventional microscopy, but still lags behind modern STED or STORM setups,
whose resolution can reach 20–30 nm. We addressed this issue here by introducing
an improved gel recipe that enables an expansion factor of ~10× in each dimension,
which corresponds to an expansion of the sample volume by more than 1,000‐fold.
Our protocol, which we termed X10 microscopy, achieves a resolution of 25–30 nm
on conventional epifluorescence microscopes. X10 provides multi‐color images similar
or even superior to those produced with more challenging methods, such as STED,
STORM, and iterative expansion microscopy (iExM). X10 is therefore the cheapest
and easiest option for high‐quality super‐resolution imaging currently available.
X10 should be usable in any laboratory, irrespective of the machinery owned or
of the technical knowledge.
article_number: e45836
article_processing_charge: No
author:
- first_name: Sven M
full_name: Truckenbrodt, Sven M
id: 45812BD4-F248-11E8-B48F-1D18A9856A87
last_name: Truckenbrodt
- first_name: Manuel
full_name: Maidorn, Manuel
last_name: Maidorn
- first_name: Dagmar
full_name: Crzan, Dagmar
last_name: Crzan
- first_name: Hanna
full_name: Wildhagen, Hanna
last_name: Wildhagen
- first_name: Selda
full_name: Kabatas, Selda
last_name: Kabatas
- first_name: Silvio O
full_name: Rizzoli, Silvio O
last_name: Rizzoli
citation:
ama: Truckenbrodt SM, Maidorn M, Crzan D, Wildhagen H, Kabatas S, Rizzoli SO. X10
expansion microscopy enables 25‐nm resolution on conventional microscopes. EMBO
reports. 2018;19(9). doi:10.15252/embr.201845836
apa: Truckenbrodt, S. M., Maidorn, M., Crzan, D., Wildhagen, H., Kabatas, S., &
Rizzoli, S. O. (2018). X10 expansion microscopy enables 25‐nm resolution on conventional
microscopes. EMBO Reports. EMBO. https://doi.org/10.15252/embr.201845836
chicago: Truckenbrodt, Sven M, Manuel Maidorn, Dagmar Crzan, Hanna Wildhagen, Selda
Kabatas, and Silvio O Rizzoli. “X10 Expansion Microscopy Enables 25‐nm Resolution
on Conventional Microscopes.” EMBO Reports. EMBO, 2018. https://doi.org/10.15252/embr.201845836.
ieee: S. M. Truckenbrodt, M. Maidorn, D. Crzan, H. Wildhagen, S. Kabatas, and S.
O. Rizzoli, “X10 expansion microscopy enables 25‐nm resolution on conventional
microscopes,” EMBO reports, vol. 19, no. 9. EMBO, 2018.
ista: Truckenbrodt SM, Maidorn M, Crzan D, Wildhagen H, Kabatas S, Rizzoli SO. 2018.
X10 expansion microscopy enables 25‐nm resolution on conventional microscopes.
EMBO reports. 19(9), e45836.
mla: Truckenbrodt, Sven M., et al. “X10 Expansion Microscopy Enables 25‐nm Resolution
on Conventional Microscopes.” EMBO Reports, vol. 19, no. 9, e45836, EMBO,
2018, doi:10.15252/embr.201845836.
short: S.M. Truckenbrodt, M. Maidorn, D. Crzan, H. Wildhagen, S. Kabatas, S.O. Rizzoli,
EMBO Reports 19 (2018).
date_created: 2019-05-28T13:16:08Z
date_published: 2018-09-01T00:00:00Z
date_updated: 2023-09-19T14:52:32Z
day: '01'
ddc:
- '580'
department:
- _id: JoDa
doi: 10.15252/embr.201845836
external_id:
isi:
- '000443682200009'
file:
- access_level: open_access
checksum: 6ec90abc637f09cca3a7b6424d7e7a26
content_type: application/pdf
creator: kschuh
date_created: 2019-05-28T13:17:19Z
date_updated: 2020-07-14T12:47:32Z
file_id: '6500'
file_name: 2018_embo_Truckenbrodt.pdf
file_size: 2005572
relation: main_file
file_date_updated: 2020-07-14T12:47:32Z
has_accepted_license: '1'
intvolume: ' 19'
isi: 1
issue: '9'
language:
- iso: eng
month: '09'
oa: 1
oa_version: Published Version
publication: EMBO reports
publication_identifier:
eissn:
- 1469-3178
issn:
- 1469-221X
publication_status: published
publisher: EMBO
quality_controlled: '1'
scopus_import: '1'
status: public
title: X10 expansion microscopy enables 25‐nm resolution on conventional microscopes
tmp:
image: /images/cc_by.png
legal_code_url: https://creativecommons.org/licenses/by/4.0/legalcode
name: Creative Commons Attribution 4.0 International Public License (CC-BY 4.0)
short: CC BY (4.0)
type: journal_article
user_id: c635000d-4b10-11ee-a964-aac5a93f6ac1
volume: 19
year: '2018'
...