Detecting Caspase Activity in Drosophila Larval Imaginal Discs
Caspases are a highly specialized class of cell death proteases. Since they are synthesized as inactive full-length zymogens, activation—at least of effector caspases and to some extent also of initiator caspases—requires a proteolytic cleavage event, generating a large and a small subunit, two of each forming the active caspase. The proteolytic cleavage event generates neo-epitopes at both the C-terminus of the large subunit and the N-terminus of the small subunit. The cleaved Caspase-3 (CC3) antibody was raised against the neo-epitope of the large subunit and thus detects only cleaved, but not full-length, Caspase-3. Although raised against human cleaved Caspase-3, the CC3 antibody cross-reacts in other species and detects cleaved caspases, most notably DrICE and Dcp-1, in Drosophila. This protocol describes the procedure for use of the CC3 antibody to detect caspase activity in larval imaginal discs in Drosophila.
Key wordsDrosophila Cleaved-Caspase-3 Dronc DrICE Dcp-1 Cell death Non-apoptotic function Imaginal Disc Immunolabeling
We would like to thank Yun Fan, Ernesto Perez, and Jillian Lindblad for their technical expertise and review of the manuscript. This work was supported by grants from the National Institutes of Health (GM068016 and GM107789).
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