I am going to grow mouse BMSCs in-vitro and I want to differentiate them into osteoclasts. I am very new to this so I have been looking around for various protocols and publications for this kind of assay and I have found a good one that suggests alpha-MEM (with 10%FBS) + MCSF (50ng/ml) + RANKL (50ng/ml). Now I am looking for suppliers of alpha-MEM like sigma and Gibco but they have so many varieties that I am confused which one should be ordered. They have alpha-MEM with and without L-glutamine, also with and without ribonucleosides and nucleosides. Can someone mention what these elements are required for the assay and which one should fit my needs?
Thanks for your response Jorg. If I flush the marrow cavity of a long bone, I think it will give HSC. Would that be good for studying OCL?
Hello have you succeed in?
I have a protocol flushing the bone marrow and culture the cells with alpha MEM with L-glutamine and with ribonucleosides+ FBS 5% + RANKL 50ng/ml + MCSF 25ng/ml
It worked once ... but not anymore don't now why...
any tips?
I guess treating the cells with 5% horse serum (in DMEM plus antibiotics) for 5-7 days will promote for differentiation. Also, try by treating with BMP2 (50ng/mL).
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