Journal article
A practical solution for preserving single cells for RNA sequencing.
- Abstract:
- The design and implementation of single-cell experiments is often limited by their requirement for fresh starting material. We have adapted a method for histological tissue fixation using dithio-bis(succinimidyl propionate) (DSP), or Lomant's Reagent, to stabilise cell samples for single-cell transcriptomic applications. DSP is a reversible cross-linker of free amine groups that has previously been shown to preserve tissue integrity for histology while maintaining RNA integrity and yield in bulk RNA extractions. Although RNA-seq data from DSP-fixed single cells appears to be prone to characteristic artefacts, such as slightly reduced yield of cDNA and a detectable 3' bias in comparison with fresh cells, cell preservation using DSP does not appear to substantially reduce RNA complexity at the gene level. In addition, there is evidence that instantaneous fixation of cells can reduce inter-cell technical variability. The ability of DSP-fixed cells to retain commonly used dyes, such as propidium iodide, enables the tracking of experimental sub-populations and the recording of cell viability at the point of fixation. Preserving cells using DSP will remove several barriers in the staging of single-cell experiments, including the transport of samples and the scheduling of shared equipment for downstream single-cell isolation and processing.
- Publication status:
- Published
- Peer review status:
- Peer reviewed
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- Files:
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(Preview, Version of record, pdf, 7.9MB, Terms of use)
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- Publisher copy:
- 10.1038/s41598-018-20372-7
Authors
- Publisher:
- Nature Publishing Group
- Journal:
- Scientific Reports More from this journal
- Volume:
- 8
- Issue:
- 1
- Pages:
- 2151
- Publication date:
- 2018-02-01
- Acceptance date:
- 2018-01-04
- DOI:
- EISSN:
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2045-2322
- Pmid:
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29391536
- Language:
-
English
- Keywords:
- Pubs id:
-
pubs:822775
- UUID:
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uuid:77cf5a1d-534d-4e6a-b0fd-5b42b1737e7a
- Local pid:
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pubs:822775
- Deposit date:
-
2018-02-05
- ARK identifier:
Terms of use
- Copyright holder:
- Bowden et al
- Copyright date:
- 2018
- Notes:
- © The Author(s) 2018. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
- Licence:
- CC Attribution (CC BY)
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