Overlay histogram showing U2OS cells stained with ab196445 (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 (ab196445, 1/500 dilution) for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) Alexa Fluor® 488 used at the same concentration and conditions as the primary antibody. 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.
![ab196445 staining Osteopontin in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab196445 at a working dilution of 1 in 100 (shown in green) and ab195889, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 594, shown in red) at 2µg/ml overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).](http://www.bioprodhub.com/system/product_images/ab_products/2/sub_4/4883_ab196445-234445-ab196445-ap2190127-10ug-HeLam-RETAKE.jpg)
ab196445 staining Osteopontin in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized in 0.1% Triton X-100 for 5 minutes and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated with ab196445 at a working dilution of 1 in 100 (shown in green) and ab195889, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 594, shown in red) at 2µg/ml overnight at +4°C. Nuclear DNA was labelled in blue with DAPI.Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).