Assaying proliferation and differentiation capacity of stem cells using disaggregated adult mouse epidermis

TY - JOUR

T1 - Assaying proliferation and differentiation capacity of stem cells using disaggregated adult mouse epidermis

AU - Jensen, Kim B

AU - Driskell, Ryan R

AU - Watt, Fiona M

PY - 2010/4/22

Y1 - 2010/4/22

N2 - In this protocol, we describe how to isolate keratinocytes from adult mouse epidermis, fractionate them into different sub-populations on the basis of cell surface markers and examine their function in an in vivo skin reconstitution assay with disaggregated neonatal dermal cells. We also describe how the isolated keratinocytes can be subjected to clonal analysis in vitro and in vivo and how to enrich for hair follicle-inducing dermal papilla cells in the dermal preparation. Using these approaches, it is possible to compare the capacity of different populations of adult epidermal stem cells to proliferate and to generate progeny that differentiate along the different epidermal lineages. Isolating, fractionating and grafting cells for the skin reconstitution assay is normally spread over 2 d. Clonal growth in culture is assessed after 14 d, while evaluation of the grafts is carried out after 4-5 weeks.

AB - In this protocol, we describe how to isolate keratinocytes from adult mouse epidermis, fractionate them into different sub-populations on the basis of cell surface markers and examine their function in an in vivo skin reconstitution assay with disaggregated neonatal dermal cells. We also describe how the isolated keratinocytes can be subjected to clonal analysis in vitro and in vivo and how to enrich for hair follicle-inducing dermal papilla cells in the dermal preparation. Using these approaches, it is possible to compare the capacity of different populations of adult epidermal stem cells to proliferate and to generate progeny that differentiate along the different epidermal lineages. Isolating, fractionating and grafting cells for the skin reconstitution assay is normally spread over 2 d. Clonal growth in culture is assessed after 14 d, while evaluation of the grafts is carried out after 4-5 weeks.

KW - Adult Stem Cells

KW - Animals

KW - Cells, Cultured

KW - Epidermis

KW - Flow Cytometry

KW - Mice

U2 - 10.1038/nprot.2010.39

DO - 10.1038/nprot.2010.39

M3 - Journal article

C2 - 20431535

SN - 1750-2799

VL - 5

SP - 898

EP - 911

JO - Nature Protocols (Online)

JF - Nature Protocols (Online)

IS - 5

ER -