Western blot analysis of Jurkat Apoptosis Cell Extracts #2043 using Caspase-7 Antibody, #9492.
Western blot analysis of Jurkat Apoptosis Cell Extracts #2043 using PARP Antibody #9542 (left), and Cleaved PARP (Asp214) (D64E10) XP® Rabbit mAb #5625 (right).
Western blot analysis of Jurkat Apoptosis Cell Extracts #2043 using Caspase-3 (8G10) Rabbit mAb, #9665.
Boil for 3 minutes prior to use. Load 10 µl of untreated and etoposide treated Jurkat Apoptosis Control Cell Extracts (etoposide) per lane.
Store at -20°C. Supplied in SDS Sample Buffer: 62.5 mM Tris-HCl (pH 6.8 at 25°C), 2% w/v SDS, 10% glycerol, 50 mM DTT, 0.01% w/v bromophenol blue or phenol red.Store at –20°C, or at –80°C for long-term storage.
Apoptosis Cell Extracts (Jurkat Untreated): Total cell extracts from Jurkat cells serve as a negative control. Supplied in SDS Sample Buffer.
Apoptosis Cell Extracts (Jurkat + Etoposide): Total cell extracts from Jurkat cells treated with 25 μM etoposide for 5 hours serve as a positive control for activated apoptotic cascades. Etoposide treatment induces proteolytic cleavage of various apoptosis-related proteins including caspases, IAP, and PARP. Supplied in SDS Sample Buffer.
Apoptosis is a regulated physiological process leading to cell death. Caspases, a family of cysteine acid proteases, are central regulators of apoptosis. Initiator caspases (including 8, 9, 10 and 12) are closely coupled to proapoptotic signals. Once activated, these caspases cleave and activate downstream effector caspases (including 3, 6 and 7), which in turn cleave cytoskeletal and nuclear proteins like PARP, α-fodrin, DFF and lamin A, and induce apoptosis. Cytochrome c released from mitochondria is coupled to the activation of caspase-9, a key initiator caspase (1). Proapoptotic stimuli include the FasL, TNF-α, DNA damage and ER stress. Fas and TNFR activate caspases 8 and 10 (2), DNA damage leads to the activation of caspase-9 and ER stress leads to the calcium-mediated activation of caspase-12 (3). The inhibitor of apoptosis protein (IAP) family includes XIAP and survivin and functions by binding and inhibiting several caspases (4,5). Smac/Diablo, a mitochondrial protein, is released into the cytosol upon mitochondrial stress and competes with caspases for binding of IAPs. The interaction of Smac/Diablo with IAPs relieves the inhibitory effects of the IAPs on caspases (6).
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