Anti-Glutaminase antibody (ab60709) + HeLa whole cell lysate at 50 µgSecondaryGoat Anti-Mouse IgG HRP at 1/2500 dilutiondeveloped using the ECL technique
Glutaminase was immunoprecipitated using 0.5mg Hela whole cell extract, 10µg of Mouse monoclonal to Glutaminase (ab60709) 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 ab60709.Secondary: Goat polyclonal to mouse IgG light chain specific (HRP) at 1/5000 dilution.Band: 78kDa: Glutaminase.
ab60709 at 10 ug/ml staining Glutaminase in human Hella cells by Immunocytochemistry/ Immunofluorescence.
Overlay histogram showing HeLa cells stained with ab60709 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab60709, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed.