Quantification of bovine mammary stem/progenitor cells by laser scanning and flow cytometry*

Due to the presence and activity of mammary stem cells (MaSC), growth and remodelling of mammary gland during puberty and lactation cycles is possible. In this study the number of putative mammary stem/progenitor cells was examined in 20 months old non-pregnant Holstein-Friesian heifers. Cells were double-stained with fluorescent dye-conjugated antibodies against stem cell antigen-1 (Sca-1) and fibronectin type III domain containing 3B (FNDC3B), and were analysed using scanning cytometry and flow cytometry. Nuclei were counterstained with Hoechst 33342. Scanning and flow cytometry revealed 2.43±0.32% and 1±0.37% of MaSC in total cell number, respectively. Sca-1posFNDC3Bpos cells did not express estrogen receptor (ERα), confirming their undifferentiated phenotype. In conclusion, scanning cytometry is a preferable method for evaluation of the number and localization of MaSC in situ, whereas, flow cytometry with cell sorting enables further genomic and biochemical analyses of isolated cells.