CB samples from normal full-term newborn infants were collected after normal vaginal delivery with informed written consent. CD34+ cells separated using a magnetic cell-sorting system (miniMACS; Miltenyi Biotec) were cultured as previously described (Abdelouahab et al., 2017 (link)). MSCs were obtained by culture of BM from healthy donors in plastic dishes in MSC expansion medium (Miltenyi Biotech). Chemotaxis assays in response to CXCL12 were performed as previously described (Rivière et al., 1999 ).
Msc expansion medium
Manufactured by Miltenyi Biotec
MSC expansion medium is a cell culture medium designed to support the growth and expansion of mesenchymal stem cells (MSCs) in vitro. It provides the necessary nutrients and growth factors to maintain the undifferentiated state and promote proliferation of MSCs.
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2 protocols using msc expansion medium
1
Isolation and Culture of Stem Cells from Newborns
2
Mesenchymal Stem Cell Differentiation
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MSC differentiation was induced by exchanging StemMACS iPS-Brew XF medium with differentiation medium containing 20% KSR, 1% GlutaMAX, 1% NEAA, and 0.1 mM beta-mercaptoethanol supplemented with 10 μM SB431542 (Miltenyi Biotec). Fresh medium was applied every other day and after 7 days, cells were transferred in a 1:3 ratio to a non-coated tissue culture-treated plate with MSC expansion medium (Miltenyi Biotec). Fresh medium was applied daily before splitting the cells at differentiation day 14. Process control of MSC differentiation was performed by flow cytometry and RT-qPCR at day 21. At day 21, MSCs were differentiated to adipocytes or osteocytes using StemMACS AdipoDiff Media or StemMACS OsteoDiff Media (both Miltenyi Biotec), respectively. Fresh medium was applied every 3 days for 20 days before process control by OilRed O or Alizarin Red staining, respectively.
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