A tunable refractive index matching medium for live imaging cells, tissues and model organisms
- Max Planck Institute of Molecular Cell Biology and Genetics, Germany
- Max Planck Institute for the Physics of Complex Systems, Germany
- Research Institute of Molecular Pathology, Germany
Abstract
In light microscopy, refractive index mismatches between media and sample cause spherical aberrations that often limit penetration depth and resolution. Optical clearing techniques can alleviate these mismatches, but they are so far limited to fixed samples. We present Iodixanol as a non-toxic medium supplement that allows refractive index matching in live specimens and thus a substantial improvement of the live-imaging of primary cell cultures, planarians, zebrafish and human cerebral organoids.
Data availability
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Data from: A tunable refractive index matching medium for live imaging cells, tissues and model organismsAvailable at Dryad Digital Repository under a CC0 Public Domain Dedication.
Article and author information
Author details
Nadine L Vastenhouw
Funding
Max-Planck-Gesellschaft (Individual research support programs)
- Tobias Boothe
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Clare M Waterman, National Institutes of Health, United States
Version history
- Received: March 28, 2017
- Accepted: July 13, 2017
- Accepted Manuscript published: July 14, 2017 (version 1)
- Version of Record published: September 4, 2017 (version 2)
- Version of Record updated: September 14, 2017 (version 3)
Copyright
© 2017, Boothe et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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A tunable refractive index matching medium for live imaging cells, tissues and model organisms
eLife 6:e27240.
https://doi.org/10.7554/eLife.27240
Further reading
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Previously we showed that 2D template matching (2DTM) can be used to localize macromolecular complexes in images recorded by cryogenic electron microscopy (cryo-EM) with high precision, even in the presence of noise and cellular background (Lucas et al., 2021; Lucas et al., 2022). Here, we show that once localized, these particles may be averaged together to generate high-resolution 3D reconstructions. However, regions included in the template may suffer from template bias, leading to inflated resolution estimates and making the interpretation of high-resolution features unreliable. We evaluate conditions that minimize template bias while retaining the benefits of high-precision localization, and we show that molecular features not present in the template can be reconstructed at high resolution from targets found by 2DTM, extending prior work at low-resolution. Moreover, we present a quantitative metric for template bias to aid the interpretation of 3D reconstructions calculated with particles localized using high-resolution templates and fine angular sampling.
