Rabbit Recombinant Monoclonal RIP3 phospho S232 + T231 antibody. Suitable for Dot, WB and reacts with Synthetic peptide, Mouse samples. Cited in 21 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Liquid
Monoclonal
Dot | WB | |
---|---|---|
Mouse | Expected | Tested |
Synthetic peptide | Tested | Not recommended |
Species | Dilution info | Notes |
---|---|---|
Species Synthetic peptide | Dilution info 1/1000 | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse | Dilution info Use at an assay dependent concentration. | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Mouse | Dilution info 1/1000 | Notes - |
Species | Dilution info | Notes |
---|---|---|
Species Synthetic peptide | Dilution info - | Notes - |
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Serine/threonine-protein kinase that activates necroptosis and apoptosis, two parallel forms of cell death (PubMed:27321907, PubMed:27746097, PubMed:27917412, PubMed:28607035, PubMed:32200799, PubMed:32296175). Necroptosis, a programmed cell death process in response to death-inducing TNF-alpha family members, is triggered by RIPK3 following activation by ZBP1 (PubMed:19590578, PubMed:22423968, PubMed:24012422, PubMed:24019532, PubMed:24095729, PubMed:24557836, PubMed:27321907, PubMed:27746097, PubMed:27819681, PubMed:27819682, PubMed:32200799, PubMed:32296175). Activated RIPK3 forms a necrosis-inducing complex and mediates phosphorylation of MLKL, promoting MLKL localization to the plasma membrane and execution of programmed necrosis characterized by calcium influx and plasma membrane damage (PubMed:24813849, PubMed:24813850, PubMed:27321907). In addition to TNF-induced necroptosis, necroptosis can also take place in the nucleus in response to orthomyxoviruses infection: following ZBP1 activation, which senses double-stranded Z-RNA structures, nuclear RIPK3 catalyzes phosphorylation and activation of MLKL, promoting disruption of the nuclear envelope and leakage of cellular DNA into the cytosol (PubMed:32200799, PubMed:32296175). Also regulates apoptosis: apoptosis depends on RIPK1, FADD and CASP8, and is independent of MLKL and RIPK3 kinase activity (PubMed:27321907). Phosphorylates RIPK1: RIPK1 and RIPK3 undergo reciprocal auto- and trans-phosphorylation (By similarity). In some cell types, also able to restrict viral replication by promoting cell death-independent responses (PubMed:30635240). In response to flavivirus infection in neurons, promotes a cell death-independent pathway that restricts viral replication: together with ZBP1, promotes a death-independent transcriptional program that modifies the cellular metabolism via up-regulation expression of the enzyme ACOD1/IRG1 and production of the metabolite itaconate (PubMed:30635240). Itaconate inhibits the activity of succinate dehydrogenase, generating a metabolic state in neurons that suppresses replication of viral genomes (PubMed:30635240). RIPK3 binds to and enhances the activity of three metabolic enzymes: GLUL, GLUD1, and PYGL (By similarity). These metabolic enzymes may eventually stimulate the tricarboxylic acid cycle and oxidative phosphorylation, which could result in enhanced ROS production (By similarity).
Rip3, Ripk3, Rip3, Receptor-interacting serine/threonine-protein kinase 3, RIP-like protein kinase 3, Receptor-interacting protein 3, RIP-3, mRIP3
Rabbit Recombinant Monoclonal RIP3 phospho S232 + T231 antibody. Suitable for Dot, WB and reacts with Synthetic peptide, Mouse samples. Cited in 21 publications.
IgG
Rabbit
pH: 7.2 - 7.4
Preservative: 0.01% Sodium azide
Constituents: PBS, 40% Glycerol (glycerin, glycerine), 0.05% BSA
Liquid
Monoclonal
EPR19403-52
Affinity purification Protein A
Blue Ice
1-2 weeks
+4°C
-20°C
Upon delivery aliquot
Avoid freeze / thaw cycle
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
This product is a recombinant monoclonal antibody, which offers several advantages including:
For more information, read more on recombinant antibodies.
This supplementary information is collated from multiple sources and compiled automatically.
'Receptor-interacting protein kinase 3' (RIP3) also known as RIPK3 is a serine/threonine-protein kinase with a molecular weight of approximately 57 kDa. Mechanically it contains a kinase domain that allows it to phosphorylate specific substrates which is important for mediating its function within the cell. RIP3 is expressed in various tissues with notable presence in the spleen heart and adipose tissue. The protein localizes predominantly in the cytoplasm where it interacts with other cellular proteins to initiate downstream signaling events.
RIP3 facilitates the execution of necroptosis a form of programmed cell death distinct from apoptosis. It becomes activated upon binding with RIP1 forming a necrosome complex that is essential for this pathway. This complex promotes phosphorylation events that subsequently lead to membrane rupture and cell death. Apart from its role in necroptosis RIP3 also engages in metabolic regulation processes linking energy status and cell death under conditions of stress.
RIP3 is a principal component of the necroptotic pathway interacting closely with RIP1 to trigger cell death in conditions where caspase activation is inhibited. Alternatively it integrates into metabolic pathways participating in sensing and responding to changes in cellular energy states. The interplay between RIP3 and RIP1 within these pathways illustrates their shared involvement in maintaining cellular homeostasis and triggering cell death when necessary.
RIP3 has significant implications for conditions involving excessive or dysfunctional cell death such as inflammatory diseases and reperfusion injury. The necroptotic activity of RIP3 can exacerbate inflammation by promoting the release of pro-inflammatory factors upon cell death. Furthermore during ischemia-reperfusion injury increased RIP3 activity in conjunction with MLKL another necroptosis-associated protein contributes to tissue damage highlighting its potential as a therapeutic target for reducing cell death-related tissue damage.
We have tested this species and application combination and it works. It is covered by our product promise.
We have not tested this specific species and application combination in-house, but expect it will work. It is covered by our product promise.
This species and application combination has not been tested, but we predict it will work based on strong homology. However, this combination is not covered by our product promise.
We do not recommend this combination. It is not covered by our product promise.
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Terms & Conditions.
Blocking/Dilution buffer: 5% NFDM/TBST.
Cells were lysed in 4% SDS buffer (4% SDS,10% 2-ME, 20% glycerol, 0.004% Bromophenol blue, Tris-HCl).
The images were kindly provided by our collaborator Dr. Jiahuai Han, Xiamen University.
All lanes: Western blot - Anti-RIP3 (phospho T231 + S232) antibody [EPR19403-52] (ab222320) at 1/500 dilution
Lane 1: Untreated L-929 (mouse connective tissue fibroblast cell line) whole cell lysate at 20 µg
Lane 2: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate treated with 10 ng/ml TNF-a and 20 µM z-VAD for 2 hours at 20 µg
Lane 3: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate treated with 10 ng/ml TNF-a and 20 µM z-VAD for 3.5 hours at 20 µg
Lane 4: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 6 hours at 20 µg
Lane 5: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate with RIP3 KO cells treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 6 hours at 20 µg
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/5000 dilution
Predicted band size: 57 kDa
Exposure time: 60s
Blocking/Dilution buffer: 5% NFDM/TBST.
The higher MW bands in Lanes 1 and 2 are extra bands.
The lysates were prepared following the 1% SDS Hot Lysate buffer method.
For Lysate preparation protocol, please refer to the protocol section of the website and/or here.
All lanes: Western blot - Anti-RIP3 (phospho T231 + S232) antibody [EPR19403-52] (ab222320) at 1/1000 dilution
Lane 1: Untreated L-929 (mouse connective tissue fibroblast cell line) whole cell lysate at 10 µg
Lane 2: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 6 hours at 10 µg
Lane 3: L-929 (mouse connective tissue fibroblast cell line) whole cell lysate treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 µM z-VAD for 6 hours, then treated with alkaline phosphatase for 1 hour at 10 µg
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 57 kDa
Observed band size: 53 kDa
Exposure time: 30s
Dot blot analysis of RIP3 (phospho T231 + S232) labeled with ab222320 at 1/1000 dilution.
Lane 1: RIP3 (phospho T231 + S232) peptide (aa229-239);
Lane 2: RIP3 (phospho T231 + S232) peptide (aa223-236);
Lane 3: RIP3 (phospho T231) peptide (aa223-239);
Lane 4: RIP3 (phospho S232) peptide (aa223-239);
Lane 5: RIP3 non-phospho peptide (aa223-239).
Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/100000 dilution was used as secondary antibody.
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.
Blocking and diluting buffer concentration: 5% NFDM/TBST.
Anti-GAPDH antibody [EPR16891] - Loading Control ab181602 GAPDH was used as a loading control at 1/1000000 dilution.
We recommend involving downstream protein p-MLKL as a control to validate the stimulation of p-RIP3.
Lanes 1 - 4: Western blot - Anti-RIP3 (phospho T231 + S232) antibody [EPR19403-52] (ab222320) at 1/1000 dilution
Lanes 5 - 8: Western blot - Anti-MLKL (phospho S345) antibody [EPR9515(2)] (Anti-MLKL (phospho S345) antibody [EPR9515(2)] ab196436) at 1/1000 dilution
Lanes 1, 3, 5 and 7: Untreated L-929 (mouse connective tissue fibroblast cell line) whole cell lysate at 20 µg
Lanes 2 and 6: L-929 (mouse connective tissue fibroblast cell line) treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 μM z-VAD for 8 hours whole cell lysate at 20 µg
Lanes 4 and 8: L-929 (mouse connective tissue fibroblast cell line) treated with 20 ng/ml TNF-a, 100 nM Smac mimetic and 20 μM z-VAD for 6 hours whole cell lysate at 20 µg
All lanes: Western blot - Goat Anti-Rabbit IgG H&L (HRP) (Goat Anti-Rabbit IgG H&L (HRP) ab97051) at 1/20000 dilution
Predicted band size: 57 kDa, 54 kDa
Observed band size: 53 kDa
Exposure time: 180s
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