ZFIN ID: ZDB-PUB-160429-1
Ex vivo tools for the clonal analysis of zebrafish hematopoiesis
Svoboda, O., Stachura, D.L., Machonova, O., Zon, L.I., Traver, D., Bartunek, P.
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.
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.