Any suggestion that the amino acids will lead to any detrimental change on differentiation of monocytes in to macrophages in conjunction with recombinant M-CSF?
To be honest I have never used them in the presence of NEAA or MEMs to differentiate them into Macrophages. They are not required. If you have enriched monocytes, they should be easy to differentiate in the presence of MCSF or GMCSF only. ( Depends on which kind of macrophages you want M1 or M2). Some people recommend autologous serum i.e. human serum but FBS also works.
Thank you for the insight!!
I have been using M-CSF alone for differentiation of cells (whole PBMCs but not pure population) for differentiation into MDMs. I have encountered a small yield of healthy macrophages, which was not the case earlier in the same protocol. Will be including autologous human serum for next set of experiments.
Any more precaution?
Best
Alok
Keep replenishing the medium with MCSF every two days. The key is pure monocytes to begin with. Alternatively a crude way is to put PBMCs in a flask with medium without serum or 0.1 % serum for an hour at 37 in the incubator. Monocytes will stick to the flask and wash away non adherent cells and replace the medium with RPMI contain serum and MCSF. Your yield will be better. This is an old technique but still works. Good luck!
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