Application
Peptide Competition and Induction: Extracts prepared from ALLN pretreated Jurkat cells left unstimulated (1, 2) or stimulated with TNF alpha?(3 7) were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were blocked with a 5% BSA-TBST buffer for one hour at room temperature and incubated with anti-IKBa pan (1, 3) or? (2, 4-7) overnight at 4?C in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1-4), the non phosphopeptide corresponding to the immunogen (5), a generic phosphoserine-containing peptide (6), or, the phosphopeptide immunogen (7). After washing, membranes were incubated with goat F(ab?)2 anti-rabbit IgG HRP conjugate and bands were detected using the Pierce SuperSignal method. The data show that the phosphorylation of IKBa is induced by the addition of TNF alpha?and that the peptide corresponding to IKBa [pSpS32/36] blocks the antibody signal, thereby demonstrating the specificity of the antibody. The data also show that treatment with TNF alpha?leads to degradation of IKBa protein (compare lanes 1 and 3), as expected, even in the presence of ALLN.| Product Name | IKB alpha (phospho Ser32/Ser36) antibody |
|---|---|
| Antibody Type | Primary Antibodies |
| Product description | Rabbit polyclonal to IKB alpha ( phospho S32 + S36 )Inhibitor of NFKBa (IKBa) is a ~40 kDa protein that has a central role in the regulation of transcription mediated by the NFKB transcription factor. In the absence of a stimulus, NFKB is sequestered in the cytoplasm through binding to IKB proteins. A variety of stimuli (e.g., proinflammatory cytokines, Tumor Necrosis Factor alpha, and interleukin-1 beta) lead to phosphorylation and activation of the IKK complex (consisting of IKK alpha, beta, and gamma). The activated IKK complex in turn phosphorylates IKBa serines 32 and 36, leading to its subsequent ubiquitination and degradation by the proteasome. The free NFKB translocates to the nucleus and stimulates transcription of NFKB-responsive genes. Phosphorylation of these serine residues by several other kinases, including p90S6 kinase (Rsk1), NFKB Inducing Kinase (NIK), and Mitogen-Activated Protein Kinase Kinase Kinase 1 (MEKK1), with subsequent activation of NFKB-mediated transcription, has also been reported. A component of the NFKB complex, RelA, has recently been shown to regulate the level of IKBa phosphorylation. |
| Immunogen | Synthetic phosphopeptide derived from the region of human IKBa that contains serines 32 and 36. |
| Modification | p-Ser32/Ser36 |
| Clonality | Polyclonal |
|---|---|
| Isotype | IgG |
| Host Species | Rabbit |
| Tested Applications | WB |
| WB: Use at a dilution of 1/1000. Detects a band of approximately 40 kDa. Optimal dilutions/concentrations should be determined by the end user.: |
|
| Species Reactivity | HumanMousePigRat |
| Concentration | 1mg/ml |
| Purification | Affinity purified |
| Gene Synonyms | IKBA MAD-3 NFKBI MAD 3 MAD3 IKB alpha (phospho S32/S36) IKB alpha (phospho Ser32/Ser36) IKB alpha (pS32/pS36) IKB alpha (pSer32/pSer36) IKB alpha phospho S32/S36 IKB alpha phospho Ser32/Ser36 Phospho IKB alpha (S32/S36) Phospho IKB alpha (Ser32 |
|---|---|
| Target | IKB alpha |
| Cellular Localization | Cytoplasmic |
Application
Peptide Competition and Induction: Extracts prepared from ALLN pretreated Jurkat cells left unstimulated (1, 2) or stimulated with TNF alpha?(3 7) were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were blocked with a 5% BSA-TBST buffer for one hour at room temperature and incubated with anti-IKBa pan (1, 3) or? (2, 4-7) overnight at 4?C in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1-4), the non phosphopeptide corresponding to the immunogen (5), a generic phosphoserine-containing peptide (6), or, the phosphopeptide immunogen (7). After washing, membranes were incubated with goat F(ab?)2 anti-rabbit IgG HRP conjugate and bands were detected using the Pierce SuperSignal method. The data show that the phosphorylation of IKBa is induced by the addition of TNF alpha?and that the peptide corresponding to IKBa [pSpS32/36] blocks the antibody signal, thereby demonstrating the specificity of the antibody. The data also show that treatment with TNF alpha?leads to degradation of IKBa protein (compare lanes 1 and 3), as expected, even in the presence of ALLN.| Positive Control | Jurkat cells pretreated for 30 minutes with N-acetyl-leucinyl-leucinyl-norleucinal (ALLN) and stimulated with TNF alpha |
|---|---|
| Application Notes | WB: Use at a dilution of 1/1000. Detects a band of approximately 40 kDa. Optimal dilutions/concentrations should be determined by the end user.: |
| Form | Liquid |
|---|---|
| Storage Instructions | Keep as concentrated solution. Store at 4°C short term. For extended storage aliquot and store at -20°C or below. Avoid freeze-thaw cycles. |
| Storage Buffer | Preservative: 0.05% Sodium Azide; Constituents: 50% Glycerol, PBS, 1mg/ml BSA. pH 7.3 |
