hello
You can use this protocol
Chondrogenic differentiation protocol
centrifuged cells in tube at 200×g for 5 min at RT. After removing the culture medium, cells were resuspended in DMEM-F12
medium and centrifuged at 200×g for 5 min at RT . After
aspirating the medium, cells were resuspended in chondrogenic
differentiation medium containing chondrogenic supplement
10 % MSCFBS, 100 μg/ml of L-glutamate, 100 μg/ml
streptomycin and 100 U/l penicillin and then centrifuged at
200×g for 5 min at RT. After centrifugation, the medium
was not removed from the tube in order not to disturb the
cell pellet. The cap was then loosened to allow gas exchange
and the tube was incubated at 37 °C in a humidified atmosphere
containing 5 % CO2 and 95 % air. The medium was
replaced with fresh medium every three days without disturbing
the cell pellet
.
Osteogenic differentiation protocol
culturing the cells induced in DMEM-LG, 10% FCS, 0.1 μM dexamethasone , 50 μM L-ascorbic acid-2-phosphate , 10mM β-glycerol phosphate disodium salt pentahydrate, and 0.3mM inorganic (sodium) phosphate .
For osteogenic differentiation, affter cell density over 90%(all most is full) I change medium plus
1 μM dexamethasone
50 μM L-ascorbic acid-2-phosphate
10mM β-glycerol phosphate
10mM BMP2
Change it every 2 day and you can analysis by staining afrer 3-4 day induction.
HI, for osteogenic differentiation we use
DMEM basal media with 10%FBS
10nm Dexamethazone
150 µg/ml L-ascorbic acid
change medium every 3 days.
for CHondrogenic differentiation
we grow the cells (min number 5x10^5 -1x10^6cells/pellet) in a falcon tube so it they will grow in a cell pellet but it has to be slightly loose to allow gas exchange initially at least and keep the tube lid slightly loose.
medium: adapted from(Lin et al., 2005, Sekiya et al., 2002)
DMEM supplemented with 10% FBS, insulin (6.25 μg/ml), dexamethasone (100 nM), ascorbate-2-phosphate (50 nM), transforming growth factor (TGF-β1, 10 ng/ml) and sodium pyruvate (110 μg/ml).
I hope that helps
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