期刊
STEM CELLS
卷 26, 期 10, 页码 2586-2594出版社
ALPHAMED PRESS
DOI: 10.1634/stemcells.2008-0498
关键词
Neural stem cells; Neurospheres; Microwell arrays; Hydrogels; Cell culture; Clonal assays
资金
- Swiss National Science Foundation [FN 205321-112323/1]
- European Young Investigator Awards
The neurosphere assay is the standard retrospective assay to test the self-renewal capability and multipotency of neural stem cells (NSCs) in vitro. However, it has recently become clear that not all neurospheres are derived from a NSC and that on conventional cell culture substrates, neurosphere motility may cause frequent neurosphere merging [ Nat Methods 2006; 3: 801-806; Stem Cells 2007; 25: 871-874]. Combining biomimetic hydrogel matrix technology with microengineering, we developed a microwell array platform on which NSC fate and neurosphere formation can be unequivocally attributed to a single founding cell. Using time-lapse microscopy and retrospective immunostaining, the fate of several hundred single NSCs was quantified. Compared with conventional neurosphere culture methods on plastic dishes, we detected a more than 100% increase in single NSC viability on soft hydrogels. Effective confinement of single proliferating cells to microwells led to neurosphere formation of vastly different sizes, a high percentage of which showed stem cell phenotypes after one week in culture. The reliability and increased throughput of this platform should help to better elucidate the function of sphere-forming stem/progenitor cells independent of their proliferation dynamics. STEM CELLS 2008; 26: 2586-2594
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