Stepwise direct HSC differentiation was performed as previously described9 (link),11 (link). Undifferentiated colonies of hPSCs were prepared at a density of <5 colonies per well. When the colonies grew to ~500 µm in diameter, they were cultured in hematopoietic differentiation medium (HDM) containing Stemline II serum-free medium (Sigma), Insulin-Transferrin-Selenium (Gibco) and BMP4 (20 and 100 ng/mL) for the first 2 days. The colonies were incubated with HDM containing 20 ng/mL BMP4 for 2 days, followed by treatment with 40 ng/mL vascular endothelial growth factor and 50 ng/mL stem cell factor (SCF) for 2 days. On day 6, the cultures were given fresh HDM supplemented with hematopoietic cocktail (50 ng/mL SCF, 10 ng/mL thrombopoietin, 50 ng/mL interleukin-3 (IL-3), 50 ng/mL granulocyte colony-stimulating factor and 50 ng/mL Feline McDonough Sarcoma (FMS)-like tyrosine kinase 3 ligand (all from R&D Systems)) and cultured for 13 days. The medium was changed every 3 days. The induction efficiency of hPSCs into the hematopoietic lineage was assessed by measuring the frequencies of hemogenic precursors (CD45-CD31+), hematopoietic progenitors (CD34+CD45+ or CD34+CD43+) and mature blood cells (CD34-CD45+ and CD34-CD43+) by flow cytometry.
Stemline 2 serum free medium
Manufactured by Merck Group
Stemline II serum-free medium is a cell culture media formulation designed to support the growth and maintenance of various cell types, including stem cells, in a serum-free environment. It provides the necessary nutrients and growth factors to sustain cell viability and proliferation without the need for animal-derived serum supplements.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using stemline 2 serum free medium
1
Stepwise Differentiation of hPSCs into Hematopoietic Cells
2
Hematopoietic Differentiation of hPSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Hematopoietic differentiation was performed as previously described (Hong et al., 2011 (link)). Briefly, undifferentiated hPSC colonies were prepared with low density (approximately 5–7 colonies in a 35 mm culture dish). Cells were grown to 1 mm size, media were changed to Stemline II serum-free medium (Sigma) supplemented with Insulin-Transferrin-Selenium and BMP4 (20 ng/ml) for 4 days, followed by treatment with SCF (50 ng/ml) and VEGF (40 ng/ml) for 2 days. On day 6, the cultures were given fresh hematopoietic induction medium supplemented with hGFs cocktail (50 ng/ml SCF, 10 ng/ml Thrombopoietin, 50 ng/ml Interleukin-3, 50 ng/ml FMS-like tyrosine kinase 3 ligand and 50 ng/ml Granulocyte colony-stimulating factor, R&D Systems) with and without 10 μM Im and cultured for 11 days. The hematopoietic differentiation was assessed by the frequencies of CD34+CD45+ and CD34−CD45+ populations on day 17.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!