This data was developed using the same antibody clone in a different buffer formulation containing PBS and sodium azide (ab6161)

ab6161 staining Tubulin in HeLa cells. The cells were fixed with 4% paraformaldehyde (10 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab6161 at 5µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150165, Goat polyclonal Secondary Antibody to Rat IgG - H&L (Alexa Fluor^® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor^® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour red). Nuclear DNA was labelled with DAPI (shown in blue).

Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

Overlay histogram showing HeLa cells stained with ab6161 (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 (ab6161, 1µg/1x10⁶ cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rat IgG (H+L) (ab98386) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rat IgG2a [aRTK2758] (ab18450, 1µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.

This data was developed using the same antibody clone in a different formulation containing PBS, Azide and Arginine (ab6161).

Cultured human macrophages were used with ab6161 at 1/1000 for immunofluorescence. Cells were fixed with cold 2% formaldehyde for 20mins.

Green staining is Alexa 568, Blue staining is DAPI stain.

This cell represents a young macrophage, the staining patterns varied as the cells aged in culture.

This data was developed using the same antibody clone in a different formulation containing PBS, Azide and Arginine (ab6161).

ICC/IF image of ab6161 stained human HepG2 cells. The cells were 4% PFA fixed (10 min), permabilised in 0.1% PBS-Tween (20 min) and incubated with the antibody (ab6161, 1µg/ml) for 1h at room temperature. 1%BSA / 10% normal goat serum / 0.3M glycine was used to block non-specific protein-protein interactions. The secondary antibody (green) was Alexa Fluor^® 488 goat anti-rat IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor^® 594 WGA was used to label plasma membranes (red). DAPI was used to stain the cell nuclei (blue). This antibody also gave a positive IF result in HeLa, HEK 293 and MCF7 cells.

This data was developed using the same antibody clone in a different formulation containing PBS, Azide and Arginine (ab6161).

ab6161 staining tubulin HeLa cells treated with anisomycin (ab120495), by ICC/IF. Increase in tubulin expression correlates with increased concentration of anisomycin as described in literature.
The cells were incubated at 37°C for 6h in media containing different concentrations of ab120495 (anisomycin) in DMSO, fixed with 4% formaldehyde for 10 minutes at room temperature and blocked with PBS containing 10% goat serum, 0.3 M glycine, 1% BSA and 0.1% tween for 2h at room temperature. Staining of the treated cells with ab6161 (5 µg/ml) was performed overnight at 4°C in PBS containing 1% BSA and 0.1% tween. A DyLight 488 goat anti-rat polyclonal antibody (ab98386) at 1/250 dilution was used as the secondary antibody. Nuclei were counterstained with DAPI and are shown in blue.

This data was developed using the same antibody clone in a different formulation containing PBS, Azide and Arginine (ab6161).