Chromatin immunoprecipitation analysis of Phospho-c-Abl (pTyr89) was performed using cross-linked chromatin from 1x10^6 HCT116 colon carcinoma cells treated with serum for 0, 15, 30, and 60 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 Phospho-c-Abl (pTyr89) monoclonal antibody (Product # MA5-14796). Chromatin aliquots from ~1 x 105 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 exon 3-3 of human Bcl-x (hBcl-x) or exon 1 of human NR4A3 (hNR4A3). 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. Schematic representations of the Bcl-x and NR4A3 loci are shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions). Regions amplified by Bcl-x and NR4A3 primers are represented by black bars. Data courtesy of the Innovators Program.
|Tested species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic phosphopeptide corresponding to residues surrounding pTyr89 of human c-Abl|
|Storage buffer||0.01M HEPES, pH 7.5, with 100µg/ml BSA, 50% glycerol, 0.15M NaCl|
|Contains||<0.02% sodium azide|
|Tested Applications||Dilution *|
|ChIP assay (ChIP)||1-3 ul|
|Western Blot (WB)||1:1000|
* 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.
It is not recommended to aliquot this antibody.
The ABL1 protooncogene encodes a cytoplasmic and nuclear protein tyrosine kinase that has been implicated in processes of cell differentiation, cell division, cell adhesion, and stress response. Activity of c-Abl protein is negatively regulated by its SH3 domain, and deletion of the SH3 domain turns ABL1 into an oncogene. The t(9;22) translocation results in the head-to-tail fusion of the BCR and ABL1 genes present in many cases of chronic myelogeneous leukemia. The DNA-binding activity of the ubiquitously expressed ABL1 tyrosine kinase is regulated by CDC2-mediated phosphorylation, suggesting a cell cycle function for ABL1. The ABL1 gene is expressed as either a 6- or 7-kb mRNA transcript, with alternatively spliced first exons spliced to the common exons 2-11.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.