PUBLICATION
Isolation and Characterization of Single Cells from Zebrafish Embryos
- Authors
- Samsa, L.A., Fleming, N., Magness, S., Qian, L., Liu, J.
- ID
- ZDB-PUB-160330-5
- Date
- 2016
- Source
- Journal of visualized experiments : JoVE (109): (Journal)
- Registered Authors
- Fleming, Nicole, Liu, Jiandong, Samsa, Leigh Ann
- Keywords
- none
- MeSH Terms
-
- Animals
- Cell Separation/methods*
- Cells, Cultured
- Embryo, Nonmammalian/cytology
- Primary Cell Culture/methods*
- Transcriptome
- Zebrafish/embryology*
- Zebrafish/metabolism
- PubMed
- 27022828 Full text @ J. Vis. Exp.
Citation
Samsa, L.A., Fleming, N., Magness, S., Qian, L., Liu, J. (2016) Isolation and Characterization of Single Cells from Zebrafish Embryos. Journal of visualized experiments : JoVE. (109).
Abstract
The zebrafish (Danio rerio) is a powerful model organism to study vertebrate development. Though many aspects of zebrafish embryonic development have been described at the morphological level, little is known about the molecular basis of cellular changes that occur as the organism develops. With recent advancements in microfluidics and multiplexing technologies, it is now possible to characterize gene expression in single cells. This allows for investigation of heterogeneity between individual cells of specific cell populations to identify and classify cell subtypes, characterize intermediate states that occur during cell differentiation, and explore differential cellular responses to stimuli. This study describes a protocol to isolate viable, single cells from zebrafish embryos for high throughput multiplexing assays. This method may be rapidly applied to any zebrafish embryonic cell type with fluorescent markers. An extension of this method may also be used in combination with high throughput sequencing technologies to fully characterize the transcriptome of single cells. As proof of principle, the relative abundance of cardiac differentiation markers was assessed in isolated, single cells derived from nkx2.5 positive cardiac progenitors. By evaluation of gene expression at the single cell level and at a single time point, the data support a model in which cardiac progenitors coexist with differentiating progeny. The method and work flow described here is broadly applicable to the zebrafish research community, requiring only a labeled transgenic fish line and access to microfluidics technologies.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping