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Question: I would like to use FAM-FLICA-DEVD-FMK (for caspase 3/7, in FAM) and the 660-FLICA-YVAD-FMK (for caspase 1, in 660), in the same flow cytometry panel. I was wondering if this is possible or if it will lead to technical issues.
In addition, I would like to know if it is possible to permeabilize the cells (after having incubated the cells with FLICA), in order to stain intracellular molecules, such as transcription factors.
Answer: The FAM FLICA Caspase-3/7 Assay (for caspase-3/7, in FAM) and the FLICA 660 Caspase-1 Assay (for caspase-1, in 660) have extremely well separated excitation emission profiles and there should be no issue using these two different reagents in the same panel.
As for the question on permeablizing the cells: after having incubated the cells with FLICA you can definitely permeabilize and fix your cells for intracellular antibody staining, in order to stain intracellular molecules, such as transcription factors. The FLICA reagents bind irreversibly to the active sites of the caspases and because they are covalently bound to the caspase targets they cannot be washed out of the cells after binding. However, for fixation you will want to use paraformaldehyde not methanol/ethanol.
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