I'm recently buying some anti-CD3 and anti-CD28 to activate T cells. I also notice some of this clones have been conjugated with fluorescence for the purpose of labeling T cells for flow cytometry.
One thing i'm very interested is that once the conjugated antibody is binding to T cells, they will first activate T cells, which will definitely affect the result of the original T cells status (which is the status you want to check).
It reminds me of quantum physics. If you observe it, you change it.
I'm sure a lot of people never care about the effects of labeling T cells. How to ensure a marker antibody won't affect T cells functions?
Well, there are agonist and antagonist antibodies available, so for sure, they will affect cell functionality. As far as the "regular" antibodies go, I think no one has the knowledge if the supplier has tested their effect on cellular activities. You should do an mRNA profiling with each antibody you are going to use if it is a concern to you. Or you can use antagonist antibodies instead to block all the receptors and prevent the T cells from being activated.
Occssionally the manufacturers do have this information, or there may be publications using the clone for stimulation in functional assays (which essentially tells you to avoid it). As a first step, try asking the manufacturer (but you may well have to confirm this as suggested already).
If MHC-I site will not be functional then we can ensure ourselves that T-Cells are not activated with negligible affect.
It is quite possible that the antibody will affect the cells. Usually, they are applied to fixed cells for visualization purposes. On the other hand, for live cell imaging, one could use transient transfection with some fluorescent proteins (if applicable)
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