PUBLICATION

Ex vivo tools for the clonal analysis of zebrafish hematopoiesis

Authors
Svoboda, O., Stachura, D.L., Machonova, O., Zon, L.I., Traver, D., Bartunek, P.
ID
ZDB-PUB-160429-1
Date
2016
Source
Nature Protocols   11: 1007-1020 (Journal)
Registered Authors
Bartunek, Petr, Machonova, Olga, Svoboda, Ondrej, Traver, David, Zon, Leonard I.
Keywords
Cell culture, Haematopoietic stem cells, Zebrafish
MeSH Terms
  • Animals
  • Carps/blood
  • Cell Culture Techniques
  • Culture Media/chemistry
  • Cytokines/genetics*
  • Gene Expression Profiling
  • Hematopoiesis*
  • Hematopoietic Stem Cells/cytology*
  • Molecular Biology/methods*
  • Recombinant Proteins/genetics
  • Zebrafish/blood*
  • Zebrafish Proteins/genetics
PubMed
27123951 Full text @ Nat. Protoc.
Abstract
This protocol describes the ex vivo characterization of zebrafish hematopoietic progenitors. We show how to isolate zebrafish hematopoietic cells for cultivation and differentiation in colony assays in semi-solid media. We also describe procedures for the generation of recombinant zebrafish cytokines and for the isolation of carp serum, which are essential components of the medium required to grow zebrafish hematopoietic cells ex vivo. The outcome of these clonal assays can easily be evaluated using standard microscopy techniques after 3-10 d in culture. In addition, we describe how to isolate individual colonies for further imaging and gene expression profiling. In other vertebrate model organisms, ex vivo assays have been crucial for elucidating the relationships among hematopoietic stem cells (HSCs), progenitor cells and their mature progeny. The present protocol should facilitate such studies on cells derived from zebrafish.
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