Anti-Proteasome 19S S5A/ASF antibody [AH1.1]

• Flow Cyt

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Flow Cytometry - Anti-Proteasome 19S S5A/ASF antibody [AH1.1] (AB20239)

Overlay histogram showing HeLa cells stained with ab20239 (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 (ab20239, 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 IgG1 [ICIGG1] (ab91353, 2μg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

• IP

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Immunoprecipitation - Anti-Proteasome 19S S5A/ASF antibody [AH1.1] (AB20239)

Proteasome 19S S5A/ASF was immunoprecipitated using 0.5mg Hela whole cell extract, 5μg of Mouse monoclonal to Proteasome 19S S5A/ASF 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 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 ab20239.

Secondary : Goat polyclonal to mouse IgG light chain specific (HRP) at 1/20,000 dilution.

Band : 41kDa; Proteasome 19S S5A/ASF

All lanes:

Immunoprecipitation - Anti-Proteasome 19S S5A/ASF antibody [AH1.1] (ab20239)

Predicted band size: 40 kDa

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Exposure time: 20min