Abstract
Expansion microscopy (ExM) is a powerful technique to overcome the diffraction limit of light microscopy that can be applied in both tissues and cells. In ExM, samples are embedded in a swellable polymer gel to physically expand the sample and isotropically increase resolution in x, y, and z. The maximum resolution increase is limited by the expansion factor of the gel, which is four-fold for the original ExM protocol. Variations on the original ExM method have been reported that allow for greater expansion factors but at the cost of ease of adoption or versatility. Here, we systematically explore the ExM recipe space and present a novel method termed Ten-fold Robust Expansion Microscopy (TREx) that, like the original ExM method, requires no specialized equipment or procedures. We demonstrate that TREx gels expand 10-fold, can be handled easily, and can be applied to both thick mouse brain tissue sections and cultured human cells enabling high-resolution subcellular imaging with a single expansion step. Furthermore, we show that TREx can provide ultra-structural context to subcellular protein localization by combining antibody-stained samples with off-the-shelf small-molecule stains for both total protein and membranes.
| Original language | English |
|---|---|
| Article number | e73775 |
| Pages (from-to) | 1-24 |
| Number of pages | 24 |
| Journal | eLife |
| Volume | 11 |
| DOIs | |
| Publication status | Published - 18 Feb 2022 |
Bibliographical note
Funding Information:We are grateful to Sven van IJzendoorn (UMCG) and Wilco Nijenhuis (UU) for providing the Caco2 monolayer samples. We thank the Janelia light microscopy core facility for the use of confocal and lightsheet microscopes. We thank the Janelia cell culture core facility for maintaining and providing cultured cells. We thank the Janelia histology core facility for providing tissue slices. AA was supported by the Netherlands Organization for Scientific Research Spinoza Prize. LCK was supported by the European Research Council (ERC Consolidator Grant 819219). BM, ME, and PWT were supported by the Howard Hughes Medical Institute (HHMI).
Publisher Copyright:
© Damstra et al.
Funding
We are grateful to Sven van IJzendoorn (UMCG) and Wilco Nijenhuis (UU) for providing the Caco2 monolayer samples. We thank the Janelia light microscopy core facility for the use of confocal and lightsheet microscopes. We thank the Janelia cell culture core facility for maintaining and providing cultured cells. We thank the Janelia histology core facility for providing tissue slices. AA was supported by the Netherlands Organization for Scientific Research Spinoza Prize. LCK was supported by the European Research Council (ERC Consolidator Grant 819219). BM, ME, and PWT were supported by the Howard Hughes Medical Institute (HHMI).
Keywords
- Expansion microscopy
- Human
- Immunofluorescence
- Light microscopy
- Mouse
- Sub-organelle imaging
- Super-resolution