The ELISA kit says I have to do an acidification step to the supernatant to release any bound TGF-beta, but I wasn't sure if I need to pretreat NK cells at the time of plating to elicit a TGF-beta response. Does anybody know if this is necessary and possibly have a protocol/reference if it is necessary?
Thanks
Acidification is needed to expose the target epitope for the ELISA assay. It is only applied to the supernatant.
Be careful,
Acidificatiion release TGFbeta from latent complex. If you measure control and acidified supernatant you will have both, free and total TGFbeta produce by cells. Also measure control medium if you use serum suplemented medium to know the background levels, With this data you can also know latent TGFbeta= Total-free.
I do not know what is your intended assay design but if you pretreat cells maybe TGFbeta production could reach saturation. What is the pretreatment you are thinking about? Is a kind of activated control group cells?
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