Clone E289 (ab213219) has been successfully conjugated by Abcam. This image was generated using Anti-STAT5a antibody [E289] (PE). Please refer to ab211686 for protocol details.

Overlay histogram showing A549 cells stained with ab211686 (red line). The cells were fixed with 4% formaldehyde (10 min) and then permeabilized with 0.1% PBS-Triton X-100 for 15 min. The cells were then incubated in 1x PBS / 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab211686, 1/500 dilution) for 30 min at 22°C.

Isotype control antibody (black line) was rabbit IgG (monoclonal) Phycoerythrin (ab209478) 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 50 mW Yellow/Green laser (561nm) and 586/15 bandpass filter.

This antibody gave a positive signal in A549 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Triton X-100 for 15 min used under the same conditions.

Clone E289 (ab213219) has been successfully conjugated by Abcam. This image was generated using Anti-STAT5a antibody [E289] (Alexa Fluor® 647). Please refer to ab194309 for protocol details.

ab194309 staining STAT5a in A549 cells. The cells were fixed with 4% formaldehyde (10 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 ab194309 at 1/100 Dilution(shown in red) and ab195887, Mouse monoclonal [DM1A] to alpha Tubulin (Alexa Fluor® 488, shown in green) 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).

ab32043 (purified) at 1:20 dilution (0.6ug) immunoprecipitating in TF-1 whole cell lysate. TF-1 (Human Erythroleukemia erythroblast) whole cell lysate 10ug
Lane 2 (+): ab32043 & TF-1 whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab32043 in TF-1 whole cell lysate
For western blotting, VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution
Blocking and diluting buffer: 5% NFDM/TBST.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

Flow Cytometry analysis of Jurkat (human acute T cell leukemia) cells labeling STAT5a with purified ab32043 at 1:100 dilution (1 μg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor^® 488) secondary antibody was used at 1:2000 dilution. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

Immunocytochemistry/ Immunofluorescence analysis of Jurkat (human T cell leukemia T lymphocyte) cells labeling STAT5a with purified ab32043 at 1:100 (1.2 μg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with Ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) 1:200 (2.5 μg/ml). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1:1000 (2 μg/ml) dilution. DAPI nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human breast tissue sections labeling STAT5a with purified ab32043 at 1:1000 dilution (0.12 μg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Tissue was counterstained with Hematoxylin. ImmunoHistoProbe one step HRP Polymer (ready to use). PBS instead of the primary antibody was used as the negative control.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

Immunocytochemistry/Immunofluorescence analysis of Jurkat cells labelling STAT5a with unpurified ab32043 at 1/500. Cells were fixed with 4% paraformaldehyde and permeabilized with 0.1% Triton X-100. ab150077, an Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/1000) was used as the secondary antibody.
Control: PBS only.
Nuclear counter stain: DAPI.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

Overlay histogram showing Jurkat cells stained with unpurified ab32043 (red line). The cells were fixed with 4% paraformaldehyde (10 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 (ab32043, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight^® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line). Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in Jurkat cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32043).

This IHC data was generated using the same anti-STAT5a antibody clone, E289, in a different buffer formulation (cat# ab32043).

Unpurified ab32043 at 1/250 dilution, staining human squamous lung carcinoma by Immunohistochemistry, paraffin-embedded tissue.