The caspase family of cysteine proteases plays a key role in apoptosis and inflammation. Caspase-3 is a key protease that is activated during the early stages of apoptosis and, like other members of the caspase family, is synthesized as an inactive pro-enzyme that is processed in cells undergoing apoptosis by self-proteolysis and/or cleavage by another protease. The processed forms of caspases consist of large (17-22 kDa) and small (10-12 kDa) subunits which associate to form an active enzyme. Active caspase-3, a marker for cells undergoing apoptosis, consists of a heterodimer of 17 and 12 kDa subunits which is derived from the 32 kDa pro-enzyme. Active caspase-3 proteolytically cleaves and activates other caspases, as well as relevant targets in the cytoplasm, e.g., D4-GDI and Bcl-2, and in the nucleus (e.g. PARP). This antibody has been reported to specifically recognize the active form of caspase-3 in human and mouse cells. It has not been reported to recognize the pro-enzyme form of caspase-3. Immunoprecipitation/Western blot analysis of caspase-3 from apoptotic and non-apoptotic cell lysates. Jurkat cells (Human T-cell leukemia, ATCC TIB-152) were left untreated (left panel) or were treated with 6 μM camptothecin for 5 hr to induce apoptosis (right panel). Immunoprecipitation: Cell lysates were immunoprecipitated with 0.25 - 2 μg/mL of the rabbit anti- active caspase-3 antibody [clone C92-605, lanes 1 and 8 (2 μg), lanes 2 and 9, (1 μg), lanes 3 and 10 (0.5 μg), and lanes 4 and 11 (0.25 μg)], 1 - 2 μg/mL of a mAb recognizing both pro and active caspase-3 (Cat. No. 610322), lanes 5 and 12 (2 μg), lanes 6 and 13 (1 μg)] or 1 μg/mL of a rabbit IgG isotype control [Jackson Immunoresearch (Cat. No. 011-00000-3), lanes 7 and 14]. Western blot: Caspase-3 was detected by western blot analysis with an antibody that recognizes both pro- (32 kD) and active (20 and 17 kD, reflecting the presence or absence of the caspase-3 pro-domain) caspase-3 [lanes 1-14]. The results also show that the rabbit anti- active caspase-3 antibody (clone C92-605) immunoprecipitated only the active form of caspase-3 (lanes 8-11) as compared to the mouse anti-human caspase-3 antibody (Cat. No. 610322) which immunoprecipitated both the pro and active forms of caspase-3 (lanes 12 and 13). Bands may be observable at ~25 kD and ~55-60 kD which represent the light and heavy chains, respectively, of IgG used for the immunoprecipitation.
This product contains Sodium azide: a POISONOUS AND HAZARDOUS SUBSTANCE which should be handled by trained staff only.
Stock
4 °C
Stockage commentaire
Store undiluted at 4°C.
Dai, Krantz: "Interferon gamma induces upregulation and activation of caspases 1, 3, and 8 to produce apoptosis in human erythroid progenitor cells." dans: Blood, Vol. 93, Issue 10, pp. 3309-16, (1999) (PubMed).
Fujita, Tsuruo: "Involvement of Bcl-2 cleavage in the acceleration of VP-16-induced U937 cell apoptosis." dans: Biochemical and biophysical research communications, Vol. 246, Issue 2, pp. 484-8, (1998) (PubMed).