CREB Phospho specific pS133 antibody

Antigen: CREB Phospho specific pS133

Clonality: Polyclonal

Host: Rabbit

Reactivity: Human, Cow (Bovine), Mouse (Murine), Rat (Rattus)

Application: Immunohistochemistry (IHC), ELISA, Western Blotting (WB), Immunoprecipitation (IP)

Catalog no.: ABIN97863

Additional Information

Immunogen: CREB phospho peptide corresponding to a region of the human protein conjugated to Keyhole Limpet Hemocyanin (KLH). Sequence information: Q-K-R-R-E-I-L-S-R-R-P-pS-Y-R-K-I-L-N-D-L-S-S-D-A-P corresponding to aa 122-147

Format: Liquid (sterile filtered)

Description: The CREB (Cyclic AMP-response-element-binding-protein 1) gene encodes a transcription factor that is a member of the leucine zipper family of DNA binding proteins. This protein binds as a homodimer to the cAMP-responsive element (CRE element TGANNTCA), an octameric palindrome. Phosphorylation by cAMP-dependent protein kinase (PKA) at serine-119 is required for interaction with DNA and phosphorylation at serine-133 allows CREB to interact with CBP (CREB binding protein) leading to interaction with RNA polymerase II. Alternate splicing of this gene results in two transcript variants encoding different isoforms.

Specificity: This product was affinity purified from monospecific antiserum by immunoaffinity purification. Antiserum was first sequentially pre-adsorbed against a control E.coli lysate, nickel-purified recombinant CREB and the non-phosphorylated form of the immunizing peptide. The resultant depleted antiserum was then purified against the phosphorylated form of the immunizing peptide. This phospho specific polyclonal antibody is specific for phosphorylated pS133 of human CREB. Reactivity with non-phosphorylated CREB is minimal. This antibody does show cross reactivity with pS133 phosphorylated CREB from mouse and rat.

Application Details

Application Notes: Recommended Dilutions: This antibody was assayed against a variety of tissues including fibroblasts and B-cell lysates and reacts specifically with CREB when phsophorylated at serine 133. Bands of 46 and 43 kDa corresponding to phosphorylated CREB are observed in western blots using the antibody at a dilution of 1:1,000 followed by reaction with Peroxidase conjugated Affinity Purified anti-Rabbit IgG [H&L] (Goat) code ABIN101990. Anti-CREB pS133 is suitable for the detection by immunoblot of phosphorylated human, mouse and rat CREB. No cross-reactivity occurs with non-phosphorylated CREB. This antibody is also reactive in immunoprecipitation experiments and by ELISA. Optimal titers for other applications should be determined by the researcher. Note: This phospho specific polyclonal antibody reacts with phosphorylated human CREB at pS133 and shows minimal reactivity by western blot with non-phosphorylated CREB and minimal reactivity by ELISA against the non-phosphorylated form of the immunizing peptide. Although not tested, this antibody is likely functional in immunohisto-chemistry, FACS, and immunoprecipitation.

Concentration: 1.0 mg/ml (by UV absorbance at 280 nm)

Buffer: Stabilizer: None. Preservative: 0.1% (w/v) Sodium Azide. Buffer: 0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2.

Storage: Store vial at -20° C prior to opening. Centrifuge product if not completely clear after standing at room temperature. Dilute only prior to immediate use. For extended storage aliquot contents and freeze at -20° C or below. Avoid cycles of freezing and thawing. Expiration date is one (1) year from date of opening. Figure 1 ­ Anti-CREB pS133 was used to detect phosphorylated CREB by immunoblot. Recombinant His-tagged human CREB was produced in E.coli and purified by metal affinity chromatography. An aliquot of purified CREB was phosphorylated in-vitro using Protein Kinase A and ATP. Immunoblot of control (-) and in-vitro phosphorylated CREB (+) was used to show that the antibody reacts specifically with the phosphorylated form. Pan reactive CREB (# ABIN97864) reacts equally with both non-phosphorylated and phosphorylated CREB (not shown). Detection occurs using a 1:1,000 dilution of antibody followed by

Research Area: Transcription Factors

Restrictions: For Research Use only

Images

Anti-CREB pS133 was used to detect phosphorylated CREB by immunoblot. Recombinant His-tagged human CREB was produced in E.coli and purified by metal affinity chromatography. An aliquot of purified CREB was phosphorylated in-vitro using Protein Kinase A and ATP. Immunoblot of control (-) and in-vitro phosphorylated CREB (+) was used to show that the antibody reacts specifically with the phosphorylated form. Pan reactive CREB (antibodies-online # 100-401- 195) reacts equally with both nonphosphorylated and phosphorylated CREB (not shown). Detection occurs using a 1:1,000 dilution of antibody followed by 1:5,000 dilution of HRP Goat-a-Rabbit IgG with visualization via ECL. Film exposure approximately 1