RRC ID 51266
Author Sassen WA, Lehne F, Russo G, Wargenau S, Dübel S, Köster RW.
Title Embryonic zebrafish primary cell culture for transfection and live cellular and subcellular imaging.
Journal Dev. Biol.
Abstract Although having great potential for live cell imaging to address numerous cell biological questions with high spatial and temporal resolution, primary cell cultures of zebrafish embryos are not widely used. We present an easy-to-use protocol for preparing primary cell cultures of 2 dpf zebrafish embryos allowing for live cell imaging of fully differentiated cells such as neurons and myocytes. We demonstrate that different cell types can be identified by morphology and expression of transgenic cell type-specific fluorescent reporters and that fluorescent cells can be sorted by flow cytometry to prepare an enriched culture. To facilitate subcellular imaging in live primary cells, we successfully tested a selection of fluorescent vital dyes. Most importantly, we demonstrate that zebrafish primary cells can be transfected efficiently with expression constructs allowing for visualizing subcellular structures with fluorescent marker proteins for time lapse imaging. We propose zebrafish primary cell culture as a versatile tool to address cell biological questions in combination with a powerful in vivo model.
Volume 430(1)
Pages 18-31
Published 2017-10-1
DOI 10.1016/j.ydbio.2017.07.014
PII S0012-1606(17)30244-0
PMID 28802829
MeSH Animals Cell Shape Cells, Cultured Electroporation Embryo, Nonmammalian / cytology* Embryo, Nonmammalian / metabolism Flow Cytometry Imaging, Three-Dimensional* Macrophages / cytology Motor Neurons / cytology Neuroglia / cytology Primary Cell Culture / methods* Purkinje Cells / cytology Staining and Labeling Subcellular Fractions / metabolism Transfection / methods* Transgenes Zebrafish / embryology*
IF 3.262
Resource
Zebrafish Tg(CM-isl1:GFP)