Recombinant Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [E83-77] to Cleaved Caspase-3
- Suitable for: WB, ICC/IF
- Knockout validated
- Reacts with: Human, Recombinant fragment
Related conjugates and formulations
Overview
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Product name
Anti-Cleaved Caspase-3 antibody [E83-77]
See all Cleaved Caspase-3 primary antibodies -
Description
Rabbit monoclonal [E83-77] to Cleaved Caspase-3 -
Host species
Rabbit -
Specificity
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information. This antibody only detects the active (cleaved) form of Caspase-3 and does not recognize the pro form of Caspase-3. PubMedID 19789217 describes the detection of human cells injected into mice. The pro-caspase-3 is cleaved only when apoptosis event occurs. So, in order to detect active Caspase-3, we strongly suggest to induce your samples into apoptotic pathway.
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Tested applications
Suitable for: WB, ICC/IFmore details
Unsuitable for: Flow Cyt,IHC-P or IP -
Species reactivity
Reacts with: Human, Recombinant fragment -
Immunogen
Synthetic peptide within Human Cleaved Caspase-3 aa 1-100 (N terminal). The exact sequence is proprietary. A synthetic peptide corresponding to residues following Ser29 of human Caspase 3 (N terminus of p17 subunit).
Database link: P42574 -
Positive control
- WB: Wild type HAP1 + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate; Jurkat cell lysate (camptothecin treated); HeLa cell lysate (staurosporine treated). ICC/IF: Hela cells (staurosporine treated); Human Vascular endothelial cells.
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General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles. -
Storage buffer
pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 49% PBS, 50% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
E83-77 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Assay kits
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Compatible Secondaries
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Isotype control
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Positive Controls
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Recombinant Protein
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab32042 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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WB | (7) |
1/500. Detects a band of approximately 17 kDa (predicted molecular weight: 32 kDa).
The pro-caspase-3 is cleaved only when apoptosis event occurs. So, in order to detect active Caspase-3, we strongly suggest to induce your samples into apoptotic pathway. |
ICC/IF | (5) |
1/100 - 1/250.
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Notes |
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WB
1/500. Detects a band of approximately 17 kDa (predicted molecular weight: 32 kDa). The pro-caspase-3 is cleaved only when apoptosis event occurs. So, in order to detect active Caspase-3, we strongly suggest to induce your samples into apoptotic pathway. |
ICC/IF
1/100 - 1/250. |
Target
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Function
Involved in the activation cascade of caspases responsible for apoptosis execution. At the onset of apoptosis it proteolytically cleaves poly(ADP-ribose) polymerase (PARP) at a '216-Asp-
-Gly-217' bond. Cleaves and activates sterol regulatory element binding proteins (SREBPs) between the basic helix-loop-helix leucine zipper domain and the membrane attachment domain. Cleaves and activates caspase-6, -7 and -9. Involved in the cleavage of huntingtin. Triggers cell adhesion in sympathetic neurons through RET cleavage. -
Tissue specificity
Highly expressed in lung, spleen, heart, liver and kidney. Moderate levels in brain and skeletal muscle, and low in testis. Also found in many cell lines, highest expression in cells of the immune system. -
Sequence similarities
Belongs to the peptidase C14A family. -
Post-translational
modificationsCleavage by granzyme B, caspase-6, caspase-8 and caspase-10 generates the two active subunits. Additional processing of the propeptides is likely due to the autocatalytic activity of the activated protease. Active heterodimers between the small subunit of caspase-7 protease and the large subunit of caspase-3 also occur and vice versa.
S-nitrosylated on its catalytic site cysteine in unstimulated human cell lines and denitrosylated upon activation of the Fas apoptotic pathway, associated with an increase in intracellular caspase activity. Fas therefore activates caspase-3 not only by inducing the cleavage of the caspase zymogen to its active subunits, but also by stimulating the denitrosylation of its active site thiol. -
Cellular localization
Cytoplasm. - Information by UniProt
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Database links
- Entrez Gene: 836 Human
- Omim: 600636 Human
- SwissProt: P42574 Human
- Unigene: 141125 Human
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Alternative names
- active caspase 3 antibody
- Apopain antibody
- CASP 3 antibody
see all
Images
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All lanes : Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution
Lane 1 : Wild-type HeLa Treated Staurosporine (2uM, 4h) cell lysate
Lane 2 : CASP3 knockout HeLa Treated Staurosporine (2uM, 4h) cell lysate
Lane 3 : Wild-type HeLa Vehicle Control Staurosporine (0uM, 4h) cell lysate
Lane 4 : CASP3 knockout HeLa Vehicle Control Staurosporine (0uM, 4h) cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 32 kDa
Observed band size: 16,28 kDa why is the actual band size different from the predicted?Western blot: Anti-CASP3 antibody [E83-77] (ab32042) staining at 1/500 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab32042 was shown to bind specifically to CASP3. A band was observed at 16/28 kDa in treated wild-type HeLa cell lysates with no signal observed at this size in CASP3 knockout cell line. To generate this image, wild-type and CASP3 knockout HeLa cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
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Lane 1: Wild type HAP1 + DMSO for 24 hours, whole cell lysate (20 µg)
Lane 2: Wild type HAP1 + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 µg)
Lane 3: HAP1 CASP3 KO + DMSO for 24 hours, whole cell lysate (20 µg)
Lane 4: HAP1 CASP3 KO + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 µg)
Lane 5: HeLa + DMSO for 24 hours, whole cell lysate (20 µg)
Lane 6: HeLa + 2uM Staurosporine (ab146588) for 24 hours, whole cell lysate (20 µg)
Lanes 1 - 6: Merged signal (red and green). Green - ab32042 observed at 17 kDa. Red - loading control, ab130007, observed at 130 kDa.ab32042 was shown to specifically react with CASP3 (Caspase-3) when CASP3 (Caspase-3) knockout samples were used. HAP1 wild-type and CASP3 (Caspase-3) knockout samples were subjected to SDS-PAGE. Ab32042 and ab130007 (Mouse anti vinculin loading control) were incubated overnight at 4°C at 500 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.
Cells were grown to confluency prior to treatment.
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High-glucose induces apoptosis in human Vascular endothelial cells (VECs).
Apoptotic responses in VEC were analyzed by detection of cleaved-Caspase-3 immunofluorescence using ab32042. Cells were treated with low glucose (LG) or high glucose (HG) for 72 hours before treated with 100 ng/mL bFGF, 1 μM sp600125 (sp), or 1 μM U0126 (U) or 10 μM MnTmPyP for 1 hour. Bar = 100 μm.
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Ab32042, at dilution of 1/100, staining HeLa (human epithelial cell line from cervix adenocarcinoma) cells by Immunofluorescence.
Left image: control.
Right image: staurosporine treated. -
All lanes : Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution
Lane 1 : HeLa Whole Cell Lysate (2 uM Staurosporine, 4Hr) at 20 µg
Lane 2 : HeLa Whole Cell Lysate (untreated) at 20 µg
Lane 3 : Cleaved Caspase 3 (recombinant protein) at 0.1 µg
Secondary
All lanes : 800CW Goat Anti-Rabbit IgG at 1/10000 dilution
Performed under reducing conditions.
Predicted band size: 32 kDa
Additional bands at: 17 kDa (possible mature (processed) protein) -
Lane 1 : anti Pro Caspase 3 at 1/10000 dilution
Lane 2 : anti Pro Caspase 3 at 1/10000 dilution
Lanes 3-4 : Anti-Cleaved Caspase-3 antibody [E83-77] (ab32042) at 1/500 dilution
Lane 1 : Jurkat (human T cell leukemia cell line from peripheral blood) cell lysate
Lanes 2 & 4 : Jurkat cell lysate + Camptothecin
Lane 3 : Jurkat cell lysate
Predicted band size: 32 kDa
Observed band size: 17 kDa why is the actual band size different from the predicted?
Additional bands at: 30 kDa. We are unsure as to the identity of these extra bands.
Datasheets and documents
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SDS download
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Datasheet download
References (463)
ab32042 has been referenced in 463 publications.
- Liu X et al. LncRNA ZFAS1 contributes to osteosarcoma progression via miR-520b and miR-520e-mediated inhibition of RHOC signaling. Clinics (Sao Paulo) 78:100143 (2023). PubMed: 36473367
- Ma J et al. Exosome-mediated lnc-ABCA12-3 promotes proliferation and glycolysis but inhibits apoptosis by regulating the toll-like receptor 4/nuclear factor kappa-B signaling pathway in esophageal squamous cell carcinoma. Korean J Physiol Pharmacol 27:61-73 (2023). PubMed: 36575934
- Wang C et al. Long non-coding RNAs LINC00689 inhibits the apoptosis of human nucleus pulposus cells via miR-3127-5p/ATG7 axis-mediated autophagy. Open Med (Wars) 17:1821-1832 (2022). PubMed: 36475062
- Yuan Z et al. AA147 ameliorates post-cardiac arrest cerebral ischemia/reperfusion injury through the co-regulation of the ATF6 and Nrf2 signaling pathways. Front Pharmacol 13:1028002 (2022). PubMed: 36506549
- Zheng X et al. Circ_0005320 promotes oral squamous cell carcinoma tumorigenesis by sponging microRNA-486-3p and microRNA-637. Bioengineered 13:440-454 (2022). PubMed: 34967281