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Flow cytometry‐based characterization of label‐retaining stem cells following transplacental BrdU labelling
Author(s) -
Poojan Shiv,
Kumar Sushil
Publication year - 2011
Publication title -
cell biology international
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.932
H-Index - 77
eISSN - 1095-8355
pISSN - 1065-6995
DOI - 10.1042/cb20100335
Subject(s) - flow cytometry , labelling , characterization (materials science) , stem cell , transplacental , microbiology and biotechnology , chemistry , biology , biochemistry , nanotechnology , materials science , fetus , genetics , pregnancy , placenta
A method to characterize and culture stem cells from neonate mouse epidermis after transplacental BrdU (bromo‐deoxyuridine) administration is described. We have characterized stem cells by their properties viz. to retain BrdU label, adhere rapidly onto collagen—fibronectin substratum and express a specific biomarker beta‐1‐integrin. BrdU‐labelled cells (detected using monoclonal antibody) constituted a sum of 18% of the total number of cells. The ability of freshly isolated keratinocytes [LRCs (label‐retaining cells)] to bind to primary BrdU antibody or to pick up PI (propidium iodide) stain was distinguishable. Viable LRCs did not retain PI. Such cells, termed EpSC (epidermis stem cell), were PI negative and BrdU positive. EpSC constituted 6% of the total cell yield. Culture in low Ca 2+ medium and susceptibility to differentiation in the presence of high Ca 2+ levels further characterized the stem cells. This protocol is useful for studying transplacental carcinogenesis.