I'm planning to transfect T-cells with a fusion protein that is up-regulated in the presence of HIV's tat molecule, which is apparently capable of increasing the expression levels of TAR-labeled sequence by 100-fold.
I want to test for possible cytotoxic effects - is there a usual assay for cytotoxicity? and if my protein is cytotoxic, what sort of modifications might be undertaken?
Hi. Common cytotoxicity assays include the MTT assay and variants such as XTT, WST-1, etc. These measure cell viability by reduction of tetrazolium or similar salts. The reduction, or lack of reduction, of these salts is indicated to be dependent upon cell viability (as well as cell number but that should be controlled for in your experiment). Otherwise, the alamar blue assay, and the LDH assay are other alternatives. These are all gross measurements of cellular viability. If you wanted to get more detailed and look for activation of signaling pathways that are involved in cytoxicity or cell death you could always look for changes in cleavage of PARP or activation of caspase-3. These may be more stringent than necessary. I think that either the MTT, WST-1, XTT, LDH, or alamar blue assays would all be accepted as viable measurements of cytotoxicity for your purposes.
Modifications to your transfection setup would depend on what you're trying to achieve by the expression of your protein of interest.
There is also Annexin-5/PI staining to screen for Apoptosis rates.
If you have toxicity concerns, try to transfect with small amounts, or use week / inducible promoters (e.g. tamoxifen or tetracyclin dependent promoters).
Standard test for cytotoxicity is 51Cr release assay. Now, labeling of target and control cells with high and low concentrations of CFSE with subsequent analysis by FACS is becoming popular.