PUBLICATION

Culture and Transfection of Zebrafish Primary Cells

Authors
Russo, G., Lehne, F., Pose Méndez, S.M., Dübel, S., Köster, R.W., Sassen, W.A.
ID
ZDB-PUB-180904-6
Date
2018
Source
Journal of visualized experiments : JoVE   (138): (Journal)
Registered Authors
Köster, Reinhard W., Russo, Giulio
Keywords
none
MeSH Terms
  • Animals
  • Primary Cell Culture/methods*
  • Transfection
  • Zebrafish/genetics*
PubMed
30175992 Full text @ J. Vis. Exp.
Abstract
Zebrafish embryos are transparent and develop rapidly outside the mother, thus allowing for excellent in vivo imaging of dynamic biological processes in an intact and developing vertebrate. However, the detailed imaging of the morphologies of distinct cell types and subcellular structures is limited in whole mounts. Therefore, we established an efficient and easy-to-use protocol to culture live primary cells from zebrafish embryos and adult tissue. In brief, 2 dpf zebrafish embryos are dechorionated, deyolked, sterilized, and dissociated to single cells with collagenase. After a filtration step, primary cells are plated onto glass bottom dishes and cultivated for several days. Fresh cultures, as much as long term differenciated ones, can be used for high resolution confocal imaging studies. The culture contains different cell types, with striated myocytes and neurons being prominent on poly-L-lysine coating. To specifically label subcellular structures by fluorescent marker proteins, we also established an electroporation protocol which allows the transfection of plasmid DNA into different cell types, including neurons. Thus, in the presence of operator defined stimuli, complex cell behavior, and intracellular dynamics of primary zebrafish cells can be assessed with high spatial and temporal resolution. In addition, by using adult zebrafish brain, we demonstrate that the described dissociation technique, as well as the basic culturing conditions, also work for adult zebrafish tissue.
Genes / Markers
Figures
Expression
Phenotype
Mutation and Transgenics
Human Disease / Model Data
Sequence Targeting Reagents
Fish
Antibodies
Orthology
Engineered Foreign Genes
Mapping
Errata and Notes