![T-bet / Tbx21 was immunoprecipitated using 0.5mg Jurkat whole cell extract, 5µg of Rabbit polyclonal to 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, Jurkat 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 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab53174.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 53kDa; T-bet / Tbx21](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_5/7922_ab53174-215265-IPVI006ab5317420mMod.jpg)
T-bet / Tbx21 was immunoprecipitated using 0.5mg Jurkat whole cell extract, 5µg of Rabbit polyclonal to 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, Jurkat 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 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab53174.Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).Band: 53kDa; T-bet / Tbx21
All lanes : Anti-T-bet / Tbx21 antibody (ab53174) at 1 µg/mlLane 1 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell LysateLane 2 : Jurkat (Human T cell lymphoblast-like cell line) Nuclear Lysate (ab14844)Lane 3 : Jurkat Whole Cell Lysate - Staurosporine Treated (24hr, 500nM)Lane 4 : E14Tg2a (Mouse embryonic stem cell line) Whole Cell LysateLysates/proteins at 10 µg per lane.SecondaryGoat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilutionPerformed under reducing conditions.