Product Pathways - Apoptosis
Procaspase Antibody Sampler Kit #12742
|12742S||1 Kit (7 x 40 µl)||---||In Stock||---|
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|Kit Includes||Quantity||Applications||Reactivity||Homology†||MW (kDa)||Isotype|
|Caspase-3 (8G10) Rabbit mAb #9665||40 µl||W, IP||H, M, R, Mk||17, 19, 35||Rabbit IgG|
|Caspase-6 Antibody #9762||40 µl||W||H, M, R||15, 35||Rabbit|
|Caspase-7 (D2Q3L) Rabbit mAb #12827||40 µl||W||H, M, R||Mk||20, 35||Rabbit IgG|
|Caspase-8 (1C12) Mouse mAb #9746||40 µl||W, IP||H||18, 43, 57||Mouse IgG1|
|Caspase-9 (C9) Mouse mAb #9508||40 µl||W||H, M, R, Hm, Mk||47 /37/35 (H). 51 /39/37 (R,M).||Mouse IgG1|
|Lamin A/C (4C11) Mouse mAb #4777||40 µl||W, IP, IHC-P, IF-F, IF-IC, F||H, M, R, Mk||74 (Lamin A), 63 (Lamin C)||Mouse IgG2a|
|PARP Antibody #9542||40 µl||W||H, M, R, Mk||89, 116||Rabbit|
|Anti-rabbit IgG, HRP-linked Antibody #7074||100 µl||Goat|
|Anti-mouse IgG, HRP-linked Antibody #7076||100 µl||Horse|
†Species predicted to react based on 100% sequence homology.
Applications Key: W=Western Blotting, IP=Immunoprecipitation, IHC-P=Immunohistochemistry (Paraffin), IF-F=Immunofluorescence (Frozen), IF-IC=Immunofluorescence (Immunocytochemistry), F=Flow Cytometry
Reactivity Key: H=Human, M=Mouse, R=Rat, Mk=Monkey, Hm=Hamster
Western blot analysis of extracts from Jurkat and A20 cells, untreated (-) or treated with Etoposide #2200 (25 μM, overnight; +), using Caspase-7 (D2Q3L) Rabbit mAb #12827.
Western blot analysis of extracts from THP-1 cells, untreated (-) or cycloheximide-treated (CHX, 10 μg/ml, overnight; +) followed by treatment with Human Tumor Necrosis Factor-α (hTNF-α) #8902 (20 ng/ml, 4 hr; +), using Lamin A/C (4C11) Mouse mAb #4777.
Western blot analysis of extracts from Jurkat, L929, and C6 cells untreated (-) or treated with Staurosporine #9953 or cytochrome c as indicated, using Caspase 9 (C9) Mouse mAb #9508.
Western blot analysis of extracts from NIH/3T3 cells, untreated (-) or treated with Staurosporine #9953 (1 µM; +), and Jurkat cells, untreated (-) or treated with Etoposide #2200 (25 µM; +), using PARP Antibody #9542.
Western analysis of extracts from HeLa and NIH/3T3 cells, untreated (-) or treated with Staurosporine #9953 (1 μM, 3 hr; +), using Caspase-3 (8G10) Rabbit mAb #9665.
Western blot analysis of extracts from SKW6.4 cells, untreated or anti-Fas-treated (1 µg/ml), and Jurkat cells, untreated or treated with Etoposide #2200 (25 µM), using Caspase-8 (1C12) Mouse mAb #9746.
The Procaspase Antibody Sampler Kit provides an economical means to evaluate the abundance and activation of caspases. The kit contains enough primary antibody to perform at least four western blots per primary antibody.
Specificity / Sensitivity
Each antibody in the Procaspase Antibody Sampler Kit detects endogenous levels of its respective target. Caspase-3 (8G10) Rabbit mAb detects full-length (35 kDa) and the large fragment (17/19 kDa) of caspase-3 resulting from cleavage at Asp175. Caspase-6 Antibody detects both full length caspase-6 (35 kDa) and the small subunit (15 kDa) of caspase-6 resulting from cleavage at Asp193. Caspase-7 (D2Q3L) Rabbit mAb detects both the full-length (35 kDa) and the large subunit (20 kDa) of caspase-7 resulting from cleavage at Asp198. Caspase-8 (1C12) Mouse mAb detects full length (57 kDa), the cleaved intermediate p43/p41, and the p18 fragment of caspase-8. Caspase-9 (C9) Antibody detects full-length caspase-9, as well as the large fragments resulting from cleavage at Asp315 and Asp330. PARP Antibody detects full length PARP1 (116 kDa), as well as the large fragment (89 kDa) of PARP1 resulting from caspase cleavage at Asp214. Lamin A/C (4C11) Mouse mAb detects full-length lamin A and lamin C proteins, as well as the larger fragments of lamin A (50 kDa) and lamin C (41 kDa) resulting from caspase cleavage. Caspase-9 (C9) Antibody detects endogenous levels of the pro form of caspase-9 as well as cleaved fragments.
Source / Purification
Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to amino-terminal residues adjacent to (Asp175) in human caspase-3 protein, residues surrounding Pro158 of human caspase-7 protein, the carboxy-terminal sequence of the p18 fragment of human caspase-8 protein, recombinant human caspase-9 protein or human lamin A protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding the cleavage site of caspase-6 or the caspase cleavage site in PARP. Polyclonal antibodies are purified by protein A and peptide affinity chromatography.
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 2, 8, 9, 10 and 12) are closely coupled to proapoptotic signals, which include the FasL, TNF-α, and DNA damage. 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 (1,2).
Caspase-8 (FLICE, Mch5, MACH) and Caspase-9 (ICE-LAP6, Mch6) are initiator caspases. CD95 receptor (Fas/APO-1) and tumor necrosis factor receptor 1 (TNFR1) activate caspase-8, leading to the release of the caspase-8 active fragments, p18 and p10 (3-6). Cytochrome c released from the mitochondria associates with procaspase-9 (47 kDa)/Apaf 1. Apaf-1 mediated activation of caspase-9 involves intrinsic proteolytic processing resulting in cleavage at Asp315 and producing a p35 subunit. Another cleavage occurs at Asp330 producing a p37 subunit that can serve to amplify the apoptotic response (7-11).
Caspase-3 (CPP-32, Apoptain, Yama, SCA-1), Caspase-6 (Mch2), and Caspase-7 (CMH-1, Mch3, ICE-LAP3) are effector caspases (12-16). Activation of caspase-3 requires proteolytic processing of its inactive zymogen/proform into activated p17 and p12 subunits (17). Procaspase-7 is activated through proteolytic processing by upstream caspases at Asp23, Asp198, and Asp206 to produce the mature subunits (14,16). Procaspase-6 is cleaved by caspase-3 at Asp23, Asp179 and Asp193 to form active large (p18) and small (p11) subunits (7).
PARP, a 116 kDa nuclear poly (ADP-ribose) polymerase, appears to be involved in DNA repair in response to environmental stress (18). This protein can be cleaved by many ICE-like caspases in vitro (2,19) and is one of the main cleavage targets of caspase-3 in vivo (17,20). In human PARP, the cleavage occurs between Asp214 and Gly215, which separates the PARP amino-terminal DNA binding domain (24 kDa) from the carboxy-terminal catalytic domain (89 kDa) (17,19). PARP helps cells to maintain their viability; cleavage of PARP facilitates cellular disassembly and serves as a marker of cells undergoing apoptosis (21).
Lamins are nuclear membrane structural components that are important in maintaining normal cell functions such as cell cycle control, DNA replication, and chromatin organization (22-24). Lamin A/C is cleaved by caspase-6 and serves as a marker for caspase-6 activation. During apoptosis, lamin A/C is specifically cleaved into large (41-50 kDa) and small (28 kDa) fragments (24,25). The cleavage of lamins results in nuclear disregulation and cell death (26,27).
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For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology® is a trademark of Cell Signaling Technology, Inc.
Select rabbit monoclonal antibodies are developed, validated, and produced at CST using in part technology under license (granting certain rights including those under U.S. Patents No. 5,675,063 and in some instances 7,429,487) from Epitomics, Inc.