Recombinant Anti-pro Caspase-3 antibody [E83-103] (ab32499)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [E83-103] to pro Caspase-3
- Suitable for: ICC/IF, WB, IHC-P, Flow Cyt (Intra)
- Knockout validated
- Reacts with: Mouse, Human
Related conjugates and formulations
Overview
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Product name
Anti-pro Caspase-3 antibody [E83-103]
See all pro Caspase-3 primary antibodies -
Description
Rabbit monoclonal [E83-103] to pro Caspase-3 -
Host species
Rabbit -
Specificity
This antibody only detects pro-form (35kD) of caspase-3, and does not recognize any cleaved caspases.
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Tested applications
Suitable for: ICC/IF, WB, IHC-P, Flow Cyt (Intra)more details -
Species reactivity
Reacts with: Mouse, Human -
Immunogen
Synthetic peptide corresponding to Human pro Caspase-3 (N terminal).
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Positive control
- Jurkat whole cell lysate (ab7899) and human colon adenocarcinoma.
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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.
Rat: We have preliminary internal testing data to indicate this antibody may not react with this species. Please contact us for more information.
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: PBS, 50% Glycerol (glycerin, glycerine), 0.05% BSA -
Concentration information loading...
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Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
E83-103 -
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
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab32499 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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ICC/IF |
Use a concentration of 5 µg/ml.
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WB |
1/10000. Detects a band of approximately 35 kDa (predicted molecular weight: 31 kDa).
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IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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Flow Cyt (Intra) |
1/50.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
Notes |
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ICC/IF
Use a concentration of 5 µg/ml. |
WB
1/10000. Detects a band of approximately 35 kDa (predicted molecular weight: 31 kDa). |
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
Flow Cyt (Intra)
1/50. ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody. |
Target
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Relevance
Caspases are a family of cysteine proteases that are key mediators of programmed cell death or apoptosis. The precursor form of all caspases is composed of a prodomain, and large and small catalytic subunits. The active forms of caspases are generated by several stimuli including ligand-receptor interactions, growth factor deprivation and inhibitors of cellular functions. All known caspases require cleavage adjacent to aspartates to liberate one large and one small subunit, which associate into a2b2 tetramer to form the active enzyme. Gene for Caspase 3 also known as Yama, CPP32, and apopain codes for a 32-kDa protein. Caspase 3 cleaves the death substrate poly(ADP-ribose) polymerase (PARP) to a specific 85 kDa form observed during apoptosis and is inhibitable by the CrmA protein. Other Caspase 3 substrates include DNA-PK, actin, GAS2, and procaspase-6, etc. Caspase 3 is activated by cleavage events at Asp-28/Ser-29 (between N-terminal pro-domain) and Asp-175/Ser-176 (between large and small subunits) to generate a large subunit of 17-kDa and a small subunit of 12-kDa. -
Cellular localization
Cytoplasmic -
Database links
- Entrez Gene: 836 Human
- Entrez Gene: 12367 Mouse
- Omim: 600636 Human
- SwissProt: P42574 Human
- SwissProt: P70677 Mouse
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Alternative names
- CASP3 antibody
- Caspase 3 antibody
- Caspase 3 apoptosis related cysteine peptidase antibody
see all
Images
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Lane 1: Wild-type HAP1 cell lysate
Lane 2: Wild-type HAP1 cell lysate + Staurosporine (1μM for 4h)
Lane 3: Caspase-3 knockout HAP1 cell lysate
Lane 4: Caspase-3 knockout HAP1 cell lysate + Staurosporine (1μM for 4h)
Lanes 1 - 4: Merged signal (red and green). Green - ab32499 observed at 35 kDa. Red - loading control, ab8245, observed at 37 kDa.
ab32499 was shown to specifically react with pro Caspase 3 when Caspase 3 knockout samples were used. Wild-type and Caspase 3 knockout samples (± Staurosporine treatment) were subjected to SDS-PAGE. ab32499 and ab8245 (loading control to GAPDH) were diluted to 1/1000 and 1/10000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging. -
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-pro Caspase-3 antibody [E83-103] (ab32499)
Immunohistochemical analysis of paraffin-embedded human colon adenocarcinoma ab32499 at 1/250 dilution.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
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ICC/IF image of ab32499 stained HeLa cells. The cells were 4% formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32499, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899, anti-rabbit DyLight® 488 used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
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Overlay histogram showing Jurkat cells stained with ab32499 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab32499, 1/50 dilution) for 30 min at 22°C. The secondary antibody used was anti-rabbit DyLight® 488 (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.
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All lanes : Anti-pro Caspase-3 antibody [E83-103] (ab32499) at 1/10000 dilution
Lane 1 : Jurkat cell lysate
Lane 2 : Jurkat cell lysate + Camptothecin
Predicted band size: 31 kDa
Observed band size: 35 kDa why is the actual band size different from the predicted?
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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SDS download
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Datasheet download
References (66)
ab32499 has been referenced in 66 publications.
- Zhang H et al. KCNQ1OT1 regulates the retinoblastoma cell proliferation, migration and SIRT1/JNK signaling pathway by targeting miR-124/SP1 axis. Biosci Rep 41:N/A (2021). PubMed: 33345272
- Zhang Z et al. Role of lncRNA PART1 in intervertebral disc degeneration and associated underlying mechanism. Exp Ther Med 21:131 (2021). PubMed: 33376513
- Wu Y et al. Tetrandrine attenuates left ventricular dysfunction in rats with myocardial infarction. Exp Ther Med 21:119 (2021). PubMed: 33335582
- Wang D et al. Long non-coding RNA SNHG20 promotes ovarian cancer development by targeting microRNA-338-3p to regulate MCL1 expression. Oncol Lett 21:130 (2021). PubMed: 33552251
- Luo Y et al. Gypenoside XVII alleviates early diabetic retinopathy by regulating Müller cell apoptosis and autophagy in db/db mice. Eur J Pharmacol 895:173893 (2021). PubMed: 33493483