if the detected cellss were stimulated in vivo, the cellls in vitro still needed to be stimulated by specific cytokines? if needed, how long is suitable for stimulating?
We analyze GrzB expression after in vivo activation (rodent and mice) by infection directly ex vivo without any further stimulation. This works fine.
I would imagine it has to do with how long you can keep the cells (I presume you are looking at T cells) activated. If you acquire the in vivo stimulated cells and analyse expression ex vivo quick enough, no further stimulation would be required. However, if the activated cells begin to die or you are isolating a memory type cell after the in vivo stimulation ex vivo re-activation would be recommended.
We don't need in vitro activation to detect Granzyme B but for cytokines we do (my personal experiences are from human cells). However some people treat animals with Golgi stop reagents during the in vivo stimulation in order to detect cytokines directly ex vivo (and this reflects better in the in vivo Ag recognition I suppose).
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