Overlay histogram showing HeLa cells stained with ab154985 (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 (ab154985, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2 (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.
Approximately 200ug of Hela cells nuclear Extract was precipitaed with 0.5ug of ab154985 shown in lane 2(Lane 1 is 10% of Input).Western Blot analysis was carried out using ab154985 at a dilution of 1:500.
Hela cells were transfected with a JMJD3 plasmid for 2 days followed by immunostaining using ab154985 diluted 1/50.