I have problem generating plasmacytoid dendritic cells (pDC) from bone marrow of the mice. Initially, I cultured 100 million cells at 1 million cells per ml in 6 well plates. I harvested the cells after 8 days but was able to get only 10% (10 million cells) of cells from initial population. From that 10% cells, I was able to fetch only 7% pDC (700000 cells).
I use Iscoves medium supplemented with glutamine, FBS, 2-Mercaptoethanol, pencillin, Streptomycin, HEPES and murine FLT3 ligand 25ng/ml (R&D). Half of the medium was replaced with fresh medium containing FLT3 on day 3 and 5.
7 times, I could get only 10% of cells from the 8-day primary bone marrow culture.
I dont know why I am not able to get atleast 80% of cells from my culture.
Please suggest if I need to do any changes in the methods or materials.
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Plasmacytoid dendritic cells generation
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