Abstract
We here describe the methods for the isolation of distinct hematopoietic subpopulations, as defined by their immune phenotype by fluorescence-activated cell sorting, and how these cells can be analyzed even at a single-cell level for the gene expression of a number of transcription factors and other differentiation markers.
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References
Attar, R. A., Scadden, D. T. (2004) Regulation of hematopoietic stem cell growth. Leukemia 18, 1760–1768.
Payne, K. J., Crooks, G. M. (2002) Human hematopoietic lineage commitment. Immunol Rev 187, 48–64.
Zhang, Y., Payne, K. J., Zhu, Y., et al. (2005) SCL expression at critical points in human hematopoietic lineage commitment. Stem Cells 23, 852–860.
Friedman, A. D. (2007) C/EBPα induces PU.1 and interacts with AP-1 and NF-κB to regulate myeloid development. Blood Cells, Mol Dis 39, 340–343.
Ye, M., Graf, T. (2007) Early decisions in lymphoid development. Curr Opin Immunol 19, 123–128.
Kastner, P., Chan, S. (2008) PU.1: A crucial and versatile player in hematopoiesis and leukemia. Int J Biochem Cell Biol 40, 22–27.
Iwasaki, H., Somoza, C., Shigematsu, H., et al. (2005) Distinctive and indispensable roles of PU.1 in maintenance of hematopoietic stem cells and their differentiation. Blood 106, 1590–1600.
Ohnedo, K., Yamamoto, M. (2002) Roles of hematopoietic transcription factors GATA-1 and GATA-2 in the development of red blood cell lineage. Acta Haematologica 108,237–245.
Shivdasani, R. A. (2001) Molecular and transcriptional regulation of megakaryocyte differentiation. Stem Cells 19, 397–407.
Matthias, P., Rolink, A. G. (2005) Transcriptional networks in developing and mature B cells. Nat Rev Immunol 6, 497–508.
Anderson, M. K. (2006) At the crossroads: diverse roles of early thymocyte transcriptional regulators. Immunol Rev 209,191–211.
Zhu, J., Emerson, S. G. (2002) Hematopoietic cytokines, transcription factors and lineage commitment. Oncogene 21, 3295–3313.
Sitnicka, E., Buza-Vidas, N., Larsson, S., et al. (2003) Human CD34+ hematopoietic stem cells capable of multilineage engrafting NOD/SCID mice express flt3: distinct flt3 and c-kit expression and response patterns on mouse and candidate human hematopoietic stem cells. Blood 102, 881–886.
Youn, B. S., Mantel, C., Broxmeyer, H. E. (2000) Chemokines, chemokine receptors and hematopoiesis. Immunol Rev 177,150–174.
Jelkmann, W. (2004) Molecular biology of erythropoeitin. Intern Med 43,649–659.
Kaushansky, K., Drachman, J. G. (2002) Molecular and cellular biology of thrombopoietin: the primary regulator of platelet production. Oncogene 21, 3359–3367.
Miranda, M. B., Johnson, D. E. (2007) Signal transduction pathways that contribute to myeloid differentiation. Leukemia 21,1363–1377.
Weissman, I. L., Anderson, D. J., Gage, F. (2001) Stem and progenitor cells: origins, phenotypes, lineage commitments, and transdifferentiations. Annu Rev Cell Dev Biol 17, 387–403.
Manz, M. G., Miyamoto, T., Akashi, K., et al. (2002) prospective isolation of human clonogenic common myeloid progenitors. Proc Natl Acad Sci USA 99, 11872–11877.
Edvardsson, L., Dykes, J., Olsson, M. L., et al. (2004) Clonogenicity, gene expression and phenotype during neutrophil versus erythroid differentiation of cytokine-stimulated CD34+ human marrow cells in vitro. Br J Haematol 127, 451–463.
Edvardsson, L., Dykes, J., Olofsson, T. (2006) Isolation and characterization of human myeloid progenitor populations – TpoR as discriminator between common myeloid and megakaryocyte/erythroid progenitors. Exp Hematol 34, 599–609.
Adolfsson, J., Mansson, R., Buza-Vidas, N., et al. (2005) identification of Flt3+ lympho-myeloid stem cells lacking erythro-megakaryocyte potential: a revised road map for adult blood lineage commitment. Cell 91, 661–672.
Chen, L., Gao, Z., Zhu, J., et al. (2007) Identification of CD13+CD36+ cells as a common progenitor for erythroid and myeloid lineages in human bone marrow. Exp Hematol 35, 1047–1055.
Case, J., Mead, L. E., Bessler, W. K., et al. (2007) Human CD34+AC133+VEGFR-2+ cells are not endothelial progenitor cells but distinct, primitive hematopoietic progenitors. Exp Hematol 35, 1109–1118.
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© 2009 Humana Press, a part of Springer Science+Business Media, LLC
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Edvardsson, L., Olofsson, T. (2009). Real-Time PCR Analysis for Blood Cell Lineage Specific Markers. In: Bugert, P. (eds) DNA and RNA Profiling in Human Blood. METHODS IN MOLECULAR BIOLOGY™, vol 496. Humana Press. https://doi.org/10.1007/978-1-59745-553-4_21
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DOI: https://doi.org/10.1007/978-1-59745-553-4_21
Publisher Name: Humana Press
Print ISBN: 978-1-934115-93-0
Online ISBN: 978-1-59745-553-4
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