Western blot analysis of EP300 was performed with 10 µg of HeLa cells transfected with Transfection Reagent alone (Lane 1), 100nM Non-Targeting control siRNA (Lane 2), or 100nM siRNA against EP300 (Lane 3). Proteins were resolved using a NuPAGE® Novex 4-12% Bis-Tris Gel (NP0322BOX), XCell SureLock™ Electrophoresis System (EI0002), and a protein size ladder. Proteins were wet transferred to a Pierce Nitrocellulose Membrane (Product # 88025) OR Pierce PVDF Membrane (Product # 88518) and blocked with Pierce Starting Block T20 (PBS) Blocking Buffer (37539) for 1 hour at room temperature. EP300 was detected at ~ 300 kDa using EP300 Mouse monoclonal antibody (Product # MA1-16608) diluted in Pierce Starting Block T20 (PBS) Blocking Buffer 4°C overnight on a rocking platform. Pierce Goat Anti-Mouse (product # 031437) HRP-Conjugated Antibodies at a 1:2500 dilution were used and chemiluminescent detection was performed using Pierce Supersignal West Dura Maximum Sensitivity Substrate (Product # 37071). Relative density of the bands normalized to Lamin A/C (74/65 kDa). EP300 Antibody (Product # MA1-16608) confirms silencing of EP300 expression.
|Tested species reactivity||Human, Mustelid, Mouse, Non-human primate, Rat|
|Published species reactivity||Human|
|Host / Isotype||Mouse / IgG1, kappa|
|Immunogen||Fusion protein containing residues 1572-2371 of human p300.|
|Storage buffer||PBS, pH 7.4|
|Contains||0.1% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Western Blot (WB)||1:250-1:500|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
|Western Blot (WB)||See 1 publications below|
Suggested positive control: HeLa nuclear extract.
p300 is a relatively stable, ubiquitously expressed, nuclear phosphoprotein which is conserved among a variety of mammalian species. It is actively synthesized and phosphorylated in both quiescent and proliferating cells.
Cyclic AMP-responsive enhancer binding protein (CREB) binding protein (CBP) and p300 are closely related transcriptional coactivators and have been shown to directly interact with many different DNA-binding transcription factors including nuclear hormone receptors, CREB, c-Fos, C-Jun/v-Jun, c-Myb/v-Myb, TFIIB and MyoD. Both CBP and p300 have been shown to display histone acetyltransferase (HAT) activity, capable of acetylating all four core histone particles in nucleosomes. As a result of HAT activity, it has been suggested CBP and p300 may play a direct role in activating chromatin for transcription.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
Targeting of p300/CREB binding protein coactivators by simian virus 40 is mediated through p53.
MA1-16608 was used in western blot to study the role of p53 in the interaction between simian virus 40 large T antigen and human CBP/p300
|Borger DR,DeCaprio JA||Journal of virology (80:4292)||2006|