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Ex vivo tools for the clonal analysis of zebrafish hematopoiesis
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SYSNO ASEP 0473122 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Ex vivo tools for the clonal analysis of zebrafish hematopoiesis Author(s) Svoboda, Ondřej (UMG-J)
Stachura, D.L. (US)
Machoňová, Olga (UMG-J)
Zon, L.I. (US)
Traver, D. (US)
Bartůněk, Petr (UMG-J) RIDNumber of authors 6 Source Title Nature Protocols. - : Nature Publishing Group - ISSN 1754-2189
Roč. 11, č. 5 (2016), s. 1007-1020Number of pages 14 Language eng - English Country GB - United Kingdom Keywords stem-cells ; in-vitro ; vertebrate hematopoiesis ; transgenic zebrafish ; progenitor cells ; danio-rerio ; fish ; thrombopoietin ; identification ; specification Subject RIV EB - Genetics ; Molecular Biology R&D Projects LO1419 GA MŠMT - Ministry of Education, Youth and Sports (MEYS) GA16-21024S GA ČR - Czech Science Foundation (CSF) Institutional support UMG-J - RVO:68378050 UT WOS 000375154300003 DOI 10.1038/nprot.2016.053 Annotation 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. Workplace Institute of Molecular Genetics Contact Nikol Škňouřilová, nikol.sknourilova@img.cas.cz, Tel.: 241 063 217 Year of Publishing 2017
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