| Isolation of mouse mesenchymal stem cells on the basis of expression of Sca-1 and PDGFR-α. | |
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MedLine Citation:
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PMID: 23154782 Owner: NLM Status: In-Data-Review |
Abstract/OtherAbstract:
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Platelet-derived growth factor receptor α (PDGFR-α) and stem cell antigen 1 (Sca-1) have recently been identified as selective markers of mouse mesenchymal stem cells (MSCs). PDGFR-α(+)Sca-1(+) (PαS) MSCs have augmented growth potential and robust tri-lineage differentiation compared with standard culture-selected MSCs. In addition, the selective isolation of PαS MSCs avoids cellular contamination that can complicate other methods. Here we describe in detail our protocol to isolate PαS MSCs using flow cytometry. In brief, the tibia and femora are isolated and crushed using a pestle and mortar. The crushed bones are then chopped and incubated for 1 h at 37 °C in 20 ml of DMEM containing 0.2% (wt/vol) collagenase. The cell suspension is filtered before red blood cell lysis and incubated with the following antibodies: allophycocyanin (APC)-conjugated PDGFR-α, FITC-conjugated Sca-1, phycoerythrin (PE)-conjugated CD45 and Ter119. Appropriate gates are constructed on a cell sorter to exclude dead cells and lineage (CD45(+)Ter-119(+))-positive cells. Approximately 10,000 PαS MSCs may then be isolated per mouse. The total protocol takes ∼7 h to complete. |
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Authors:
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Diarmaid D Houlihan; Yo Mabuchi; Satoru Morikawa; Kunimichi Niibe; Daisuke Araki; Sadafumi Suzuki; Hideyuki Okano; Yumi Matsuzaki |
Publication Detail:
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Type: Journal Article Date: 2012-11-15 |
Journal Detail:
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Title: Nature protocols Volume: 7 ISSN: 1750-2799 ISO Abbreviation: Nat Protoc Publication Date: 2012 Nov |
Date Detail:
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Created Date: 2012-11-16 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 101284307 Medline TA: Nat Protoc Country: England |
Other Details:
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Languages: eng Pagination: 2103-11 Citation Subset: IM |
Affiliation:
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1] Centre for Liver Research, National Institute for Health Research Biomedical Research Unit, University of Birmingham, Birmingham, UK. [2]. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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