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Chromatin immunoprecipitation analysis of p300 was performed using cross-linked chromatin from 1x10^6 HCT116 colon carcinoma cells treated with serum for 0, 15, and 30 minutes. Immunoprecipitation was performed using a multiplex microplate Matrix ChIP assay (see reference for Matrix ChIP protocol: http://www.ncbi.nlm.nih.gov/pubmed/22098709) with 1.0ul/100ul well volume of a p300 polyclonal antibody (Product # PA1-848). Chromatin aliquots from ~1x10^5 cells were used per ChIP pull-down. Quantitative PCR data were done in quadruplicate using 1ul of eluted DNA in 2ul SYBR real-time PCR reactions containing primers to amplify -15kb upstream of the Egr1 gene or exon-1 or exon-2 of Egr1. PCR calibration curves were generated for each primer pair from a dilution series of sheared total genomic DNA. Quantitation of immunoprecipitated chromatin is presented as signal relative to the total amount of input chromatin. Results represent the mean +/- SEM for three experiments. A schematic representation of the Egr-1 locus is shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions), the zigzag line represents an intron, and the straight line represents upstream sequence. Regions amplified by Egr-1 primers are represented by black bars. Data courtesy of the Innovators Program.
|Tested species reactivity||Human|
|Published species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide corresponding to residues G(139) T S G P N Q G P T Q S T(151) C in the nuclear factor binding domain of human p300.|
|Purification||Antigen affinity chromatography|
|Storage buffer||PBS with 1mg/ml BSA|
|Contains||0.05% sodium azide|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|ChIP assay (ChIP)||1-3 ul|
|Western Blot (WB)||1-2 µg/ml|
* 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.
|ChIP assay (ChIP)||See 1 publications below|
PA1-848 detects p300 from human cells.
PA1-848 has been successfully used in Western blot and ChIP procedures. By Western blot, this antibody detects an ~265 kDa protein representing p300 from HeLa cell lysate.
The PA1-848 immunogen is a synthetic peptide corresponding to residues G(139) T S G P N Q G P T Q S T(151) C in the nuclear factor binding domain of human p300. The PA1-848 immunizing peptide (Cat. # PEP-053) is available for use in neutralization and control experiments.
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.
Single point mutations in CBP have been proposed as causative factors in the developmental abnormalities of Rubinstein-Taybi syndrome (RTS). Although both CBP and p300 appear to function similarly, the inability of p300 to rescue CBP malfunction in RTS suggests intrinsic functional differences between CBP and p300.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
G Protein-coupled Receptor 40 (GPR40) and Peroxisome Proliferator-activated Receptor γ (PPARγ): AN INTEGRATED TWO-RECEPTOR SIGNALING PATHWAY.
PA1-848 was used in ChIP assay to investigate GPR40 and PPARgamma signaling interactions in endothelial cells
|Wang S,Awad KS,Elinoff JM,Dougherty EJ,Ferreyra GA,Wang JY,Cai R,Sun J,Ptasinska A,Danner RL||The Journal of biological chemistry (290:19544)||2015|
E1A-associated protein p300; E1A-binding protein, 300kD; EP300; p300 HAT
EP300; KAT3B; P300; RSTS2