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Pro Caspase-3 Recombinant Rabbit Monoclonal Antibody [SR03-01]

Usd: 385

Specification

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Catalog# ET1602-39

Pro Caspase-3 Recombinant Rabbit Monoclonal Antibody [SR03-01]

Application

  • WB

  • IF-Cell

  • IF-Tissue

  • IHC-P

  • IP

  • FC

Reactivity

  • Human

  • Green monkey

BSA and Azide free
Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

Pro Caspase-3 Recombinant Rabbit Monoclonal Antibody [SR03-01]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic peptide within human Caspase-3 aa 60-100.

Species Reactivity

Human, Green monkey

Validated Applications

WB, IF-Cell, IF-Tissue, IHC-P, IP, FC

Molecular Weight

Predicted band size: 32 kDa

Positive Control

HeLa cell lysate, Jurkat cell lysate, Jurkat treated with 25μM Etoposide for 5 hours cell lysate, COS-1 cell lysate, HEK-293 cell lysate, Hela, human tonsil tissue, human spleen tissue.

Conjugation

unconjugated

Clone Number

SR03-01

RRID

AB_3069652

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). It is recommended to aliquot into single-use upon delivery. Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:2,000-1:5,000

  • IF-Cell

  • 1:50

  • IF-Tissue

  • 1:50

  • IHC-P

  • 1:50

  • FC

  • 1:20-1:50

  • IP

  • Use at an assay dependent concentration.

Target

Function

Caspase-3 is a caspase protein that interacts with caspase-8 and caspase-9. It is encoded by the CASP3 gene. CASP3 orthologs have been identified in numerous mammals for which complete genome data are available. Unique orthologs are also present in birds, lizards, lissamphibians, and teleosts. Caspase-3 shares many of the typical characteristics common to all currently-known caspases. For example, its active site contains a cysteine residue (Cys-163) and histidine residue (His-121) that stabilize the peptide bond cleavage of a protein sequence to the carboxy-terminal side of an aspartic acid when it is part of a particular 4-amino acid sequence. This specificity allows caspases to be incredibly selective, with a 20,000-fold preference for aspartic acid over glutamic acid. A key feature of caspases in the cell is that they are present as zymogens, termed procaspases, which are inactive until a biochemical change causes their activation. Each procaspase has an N-terminal large subunit of about 20 kDa followed by a smaller subunit of about 10 kDa, called p20 and p10, respectively.

Background References

1. Li H et al. Protective effect of ginsenoside Rg1 on lidocaine-induced apoptosis. Mol Med Rep 9:395-400 (2014).

2. Cejkova J et al. Suppression of alkali-induced oxidative injury in the cornea by mesenchymal stem cells growing on nanofiber scaffolds and transferred onto the damaged corneal surface. Exp Eye Res 116:312-23 (2013).

Sequence Similarity

Belongs to the peptidase C14A family.

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.

Post-translational Modification

Cleavage 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.

Subcellular Location

Cytoplasm

UNIPROT

SwissProt: P42574 Human

Synonyms

Caspase3

A830040C14Rik antibody

Apopain antibody

CASP-3 antibody

CASP3 antibody

CASP3_HUMAN antibody

Casp3a antibody

Caspase 3 antibody

Caspase 3, apoptosis-related cysteine peptidase antibody

Caspase 3, apoptosis-related cysteine protease antibody

Expand

Images

  • <span style="font-weight: bold;">☑ Cell treatment (CT),Relative expression (RE)</span><br /><br />Western blot analysis of Pro Caspase-3 on different lysates with Rabbit anti-Pro Caspase-3 antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate<br />Lane 3: Jurkat cell lysate<br />Lane 4: Jurkat treated with 25μM Etoposide for 5 hours cell lysate<br />Lane 5: MCF7 cell lysate (negative)<br />Lane 6: HEK-293 cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 32 kDa<br />Observed band size: 32 kDa<br /><br />Exposure time: 3 minutes 20 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Cell treatment (CT),Relative expression (RE)

    Western blot analysis of Pro Caspase-3 on different lysates with Rabbit anti-Pro Caspase-3 antibody (ET1602-39) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate
    Lane 3: Jurkat cell lysate
    Lane 4: Jurkat treated with 25μM Etoposide for 5 hours cell lysate
    Lane 5: MCF7 cell lysate (negative)
    Lane 6: HEK-293 cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 32 kDa
    Observed band size: 32 kDa

    Exposure time: 3 minutes 20 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1602-39) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of Pro Caspase-3 on COS-1 cell lysates with Rabbit anti-Pro Caspase-3 antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/5,000 dilution.<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 32 kDa<br />Observed band size: 32 kDa<br /><br />Exposure time: 59 seconds; ECL: K1801;<br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/5,000 dilution was used in primary antibody dilution (<a href="/products/K1803" style="font-weight: bold;text-decoration: underline;">K1803</a>) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Pro Caspase-3 on COS-1 cell lysates with Rabbit anti-Pro Caspase-3 antibody (ET1602-39) at 1/5,000 dilution.

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 32 kDa
    Observed band size: 32 kDa

    Exposure time: 59 seconds; ECL: K1801;
    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1602-39) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • <span style="font-weight: bold;">☑ Knockout (KO)</span><br /><br />All lanes: Western blot analysis of Caspase-3 with anti-Caspase-3 antibody [SR03-01] (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1:500 dilution.<br />Lane 1: Wild-type Hela whole cell lysate (20 µg).<br />Lane 2: Caspase-3 knockout Hela whole cell lysate (20 µg).<br /><br /><a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a> was shown to specifically react with Caspase-3 in wild-type Hela cells. No band was observed when Caspase-3 knockout sample was tested. Wild-type and Caspase-3 knockout samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>, 1/500) and Loading control antibody (Rabbit anti-β-actin, <a href="/products/R1207-1" style="font-weight: bold;text-decoration: underline;">R1207-1</a>, 1/1,000) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG-HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:200,000 dilution was used for 1 hour at room temperature.

    ☑ Knockout (KO)

    All lanes: Western blot analysis of Caspase-3 with anti-Caspase-3 antibody [SR03-01] (ET1602-39) at 1:500 dilution.
    Lane 1: Wild-type Hela whole cell lysate (20 µg).
    Lane 2: Caspase-3 knockout Hela whole cell lysate (20 µg).

    ET1602-39 was shown to specifically react with Caspase-3 in wild-type Hela cells. No band was observed when Caspase-3 knockout sample was tested. Wild-type and Caspase-3 knockout samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (ET1602-39, 1/500) and Loading control antibody (Rabbit anti-β-actin, R1207-1, 1/1,000) was used in 5% BSA at room temperature for 2 hours. Goat Anti-Rabbit IgG-HRP Secondary Antibody (HA1001) at 1:200,000 dilution was used for 1 hour at room temperature.

  • ICC staining of Pro Caspase-3 in Hela cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor&reg;488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

    ICC staining of Pro Caspase-3 in Hela cells (green). Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA for 15 minutes at room temperature. Cells were probed with the primary antibody (ET1602-39, 1/50) for 1 hour at room temperature, washed with PBS. Alexa Fluor®488 Goat anti-Rabbit IgG was used as the secondary antibody at 1/1,000 dilution. The nuclear counter stain is DAPI (blue).

  • Immunohistochemical analysis of paraffin-embedded human tonsil tissue using anti-Pro Caspase-3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue using anti-Pro Caspase-3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-39, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

  • Immunohistochemical analysis of paraffin-embedded human spleen tissue using anti-Pro Caspase-3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human spleen tissue using anti-Pro Caspase-3 antibody. The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 8.0-8.4) for 20 minutes.The tissues were blocked in 5% BSA for 30 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1602-39, 1/50) for 30 minutes at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"

Citation

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