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Index sorting resolves heterogeneous murine hematopoietic stem cell populations.

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Schulte, Reiner 
Wilson, Nicola K 
Prick, Janine CM 
Cossetti, Chiara 
Maj, Michal K 


Recent advances in the cellular and molecular biology of single stem cells have uncovered significant heterogeneity in the functional properties of stem cell populations. This has prompted the development of approaches to study single cells in isolation, often performed using multiparameter flow cytometry. However, many stem cell populations are too rare to test all possible cell surface marker combinations, and virtually nothing is known about functional differences associated with varying intensities of such markers. Here we describe the use of index sorting for further resolution of the flow cytometric isolation of single murine hematopoietic stem cells (HSCs). Specifically, we associate single-cell functional assay outcomes with distinct cell surface marker expression intensities. High levels of both CD150 and EPCR associate with delayed kinetics of cell division and low levels of differentiation. Moreover, cells that do not form single HSC-derived clones appear in the 7AAD(dim) fraction, suggesting that even low levels of 7AAD staining are indicative of less healthy cell populations. These data indicate that when used in combination with single-cell functional assays, index sorting is a powerful tool for refining cell isolation strategies. This approach can be broadly applied to other single-cell systems, both to improve isolation and to acquire additional cell surface marker information.



Animals, Antigens, CD, Cell Differentiation, Cell Division, Endothelial Protein C Receptor, Flow Cytometry, Hematopoietic Stem Cells, Mice, Receptors, Cell Surface, Signaling Lymphocytic Activation Molecule Family Member 1

Journal Title

Exp Hematol

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Elsevier BV
Leukaemia & Lymphoma Research (12029)
Cancer Research Uk (None)
Biotechnology and Biological Sciences Research Council (BB/I00050X/1)
Wellcome Trust (097922/Z/11/Z)
Medical Research Council (MC_PC_12009)
Leukemia & Lymphoma Society (7001-12)
Medical Research Council (MR/M008975/1)
Wellcome Trust (097922/Z/11/B)
This work was supported by grants from Leukaemia and Lymphoma Research, the Medical Research Council, the National Institute for Health Research Cambridge Biomedical Research Centre, and core support grants by the Wellcome Trust to the Cambridge Institute for Medical Research and Wellcome Trust–MRC Cambridge Stem Cell Institute. DGK is the recipient of a Canadian Institutes of Health Research Postdoctoral Fellowship and a European Hematology Association non-clinical advanced research fellowship. The authors declare that they have no conflict of interest.