Hi, I would like to irradiate OP9 stromal cells for long term co-culture with T-cell progenitors (4 - 6 weeks). Do you have a good protocol? In the literature I find levels of radiation from 2000 rad - 5000 rad -> what's optimal?
Also, do you irradiate appropriate number of cells in suspension before plating into e.g. 96-well plates or you plate them first and then irradiate? If the latter is the case, is there a need to change the medium prior to co-culturing and once the cells have adhered?
Thanks!
Hi Isabel,
I used to irradiate an appropriate number of OP9 stromal cells in suspension with 2000rad before plating into 24 well. I do this the day before the co-culture with the progenitors. Note that is better to used more 10% of the cells you need to cover the entire plate because some of them die during the process.
Good luck!
Isabel
Hi, I want to use irradiated OP9 cells too. I can't find how often it is needed to replace 0P9 cell feeder by "fresh" ones, in the context of in vitro differentiation (weeks). Any insights ?
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