Lanes 1 - 2 : Anti-CoREST antibody (ab32631) at 1 µg/mlLanes 3 - 4 : Anti-CoREST antibody (ab32631) at 1/1 dilutionLane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lane 2 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate Lane 3 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate Lane 4 : Jurkat (Human T cell lymphoblast-like cell line) Nuclear Lysate Lysates/proteins at 20 µg per lane.SecondaryGoat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilutionPerformed under reducing conditions.
PCR was performed on a SCNA1 promoter following CoRest chromatin immunoprecipition using ab32631 at a concentration of 3ug/ml from 293T cells. Lane 1 - ab24166Lane 2 - Beads only controlLane 3 - IgG only controlLane 4 - Input
![CoREST was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to CoREST and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab32631.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 65kDa: CoREST.](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_2/1005_CoREST-Primary-antibodies-ab32631-8.jpg)
CoREST was immunoprecipitated using 0.5mg Hela whole cell extract, 5µg of Rabbit polyclonal to CoREST and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, Hela whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab32631.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 65kDa: CoREST.
IHC image of CoREST staining in Human normal tonsil FFPE section, performed on a BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab32631, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX
ICC/IF image of ab32631 stained MCF7 cells. The cells were 10% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32631, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) HeLa, Hek293 and HepG2 cells at 5µg/ml.