Dexamethasone is 100nM
Glycerophosphate is 10mM
Ascorbic acid is 50ug/ml.
But I'm not understanding how to calculate the working concentration and mixing with media to prepare osteogenic media.
Dear Eliza,
Once you have supplemented your media with FBS, antibiotics and L-proline you will have a determined volume of media. Let' suppose 600 mL. Then you should weight glycerophosphate and ascorbate in separate vials. You should make the calculations attending to your desired final volume (600 mL in our example). For that calculation you will need the molecular weight of each compound. Add some medium on the vials to dissolve them quickly. They will be more concentrated now, but when you add them to the whole bottle you will achieve your desired concentrations. Next add the dexamethasone (at least in our group we use a stock solution) to one of the vials. Always calculate that the final volume will be the one of the complete bottle (in our example 600 mL). These solutions are not sterile so next you should filtrate them using a 0.22 micron filters in sterile conditions. Finally add them to the bottle of media and homogeneize. And that's it!
Hope my response has helped you. Any questions feel free to ask me!
Best regards,
Daniel
Thank you so much for your help. Could you please provide the concentration of dexamethasone and its amount to be added for example in 100ml?
Dear Eliza,
You already know the concentration of dexa (100 nM). You just need to know the initial concentration you have on your stock solution. The formula to know the volume you would need is:
Vo = (Cf x Vf) /Co
Where Vo is the initial volume you would take from your stock solution and add it to the 100 mL media, Co is the concentration of your stock solution, Cf is the final concentration (100 nM) and Vf the final volume (100 mL as you asked).
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