/wp-content/uploads/2015/09/kuperwasser-banner-500px-wide-no-bg-semi-bold.png 0 0 admin /wp-content/uploads/2015/09/kuperwasser-banner-500px-wide-no-bg-semi-bold.png admin2015-09-04 16:11:302015-09-04 16:12:52Osteoblast Differentiation Assay
Osteoblast differentiation assay
- Plate cells in culture dish (1 x 105/ 6 well plate for stromal cells). Need two wells per condition for staining, more if you want to collect RNA.
- Allow cells to grow to confluence.
- Once cells have been at confluence for 2 days begin adding differentiation media-regular growth media plus:
0.01M β-glycerol phosphate
100mM Ascorbic Acid
- Add differentiation media to one well and regular growth media to other well so that you have an undifferentiated control well of cells.
- Refeed every 2-3 days, making fresh differentiation media right before adding.
- Mineralization should occur around 17-19 days
- Stain cells treated with control and differentiation media with Alizarin Red S on day 21.
- Image cells
- Collect RNA if you set up enough wells to do so. Check changes in gene expression (these should all increase if differentiation occurred):