| Literature DB >> 17406558 |
Hideo Ema1, Yohei Morita, Satoshi Yamazaki, Azusa Matsubara, Jun Seita, Yuko Tadokoro, Hiroyoshi Kondo, Hina Takano, Hiromitsu Nakauchi.
Abstract
Mouse hematopoietic stem cells (HSCs) are the best-studied stem cells because functional assays for mouse HSCs were established earliest and purification techniques for mouse HSCs have progressed furthest. Here we describe our current protocols for the purification of CD34-/lowc-Kit+Sca-1+lineage marker- (CD34-KSL) cells, the HSC population making up approximately 0.005% of bone marrow cells in adult C557BL/6 mice. Purified HSCs have been characterized at cellular and molecular levels. Since clonal analysis is essential for the study of self-renewal and lineage commitment in HSCs, here we present our single-cell colony assay and single-cell transplantation procedures. We also introduce our immunostaining procedures for small numbers of HSCs, which are useful for signal transduction analysis. The purification of CD34-KSL cells requires approximately 6 h. Initialization of single-cell culture requires approximately 1 h. Single-cell transplantation requires approximately 6 h. Single-cell immunostaining requires approximately 2 d.Entities:
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Year: 2006 PMID： 17406558 DOI： 10.1038/nprot.2006.447
Source DB: PubMed Journal: Nat Protoc ISSN： 1750-2799 Impact factor: 13.491