I'm using z-VDVAD-FMK and z-IETD-FMK to blcok the caspase 2 and 8 activation (50uM for 7h including 1h of pretreatment). But I found both inhibitors also inhibit the caspase 3/7 activity (Caspase-Glo® 3/7 Assay, promega) which was even lower than the negative control.
Anyone has experience about these inhibitors?
Is this non-specific effect? Should I try lower working concentration or change inhibitors?
Any suggestions would be appreciated.
Hi Shujun
Both z-VDVAD-FMK and z-IETD-FMK can affect caspase 3. If you check the links below from Abcam you can see a decrease in active caspase 3 expression as concentration of Z-VDVAD-FMK and z-IETD-FMK increases.
http://www.abcam.com/z-vdvad-fmk-ab142035.html
http://www.abcam.com/z-ietd-fmk-ab141382.html
Thank you Andre,
The specificity of these chemicals make the results confused. Do you know some other inhibitors of caspase 2 and 8 have little impact on caspase 3 and 7? Or the only choice is KD or KO?
Shujun
Those are the standards inhibitors. You could try titrating z-VDVAD-FMK and z-IETD-FMK. Usually, the recommended concentration is 20uM. Maybe a smaller concentration won't affect the other caspases as much.
Hi Shujun,
ZVAD is a pan-caspase inhibitor and blocks caspase activity, so wouldn't really be useful in your work. z-IETD-FMK can be obtain in a format that only inhibits caspase-8, but I'm not sure about caspase-2. I'd contact the technical department of the company you obtained it from and ask.
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