All lanes : Anti-STAT3 antibody [EPR787Y] (ab68153) at 1/500 dilution

Lane 1 : Wild-type HAP1 whole cell lysate
Lane 2 : STAT3 knockout HAP1 whole cell lysate
Lane 3 : HeLa whole cell lysate
Lane 4 : HEK293 whole cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 88 kDa

This data was developed using the same antibody clone in a different buffer formulation (ab68153).

Lanes 1 - 4: Merged signal (red and green). Green - ab68153 observed at 92 kDa. Red - loading control, ab8245, observed at 37 kDa.

Ab68153 detected the expected band for STAT3 in wild-type cells along with additional cross-reactive bands. The band was not seen in STAT3 knockout HAP1 cells. Wild-type and STAT3 knockout samples were subjected to SDS-PAGE. Ab68153 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/500 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/10000 dilution for 1 hour at room temperature before imaging.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human brain tissue sections labelling STAT3 with unpurified ab68153 at 1/140. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).

ab68153 staining STAT3 in the human cell line HeLa (human cervix adenocarcinoma) by flow cytometry. Cells were fixed with 4% paraformaldehyde, permeabilized with 90% methanol and the sample was incubated with the primary antibody at a dilution of 1/30. A goat anti rabbit IgG (Alexa Fluor® 488) at a dilution of 1/2000 was used as the secondary antibody.

Isoytype control: Rabbit monoclonal IgG (Black)

Unlabelled 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 (ab68153).

All lanes : Anti-STAT3 antibody [EPR787Y] (ab68153) at 1/1000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : STAT3 knockout HeLa cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 88 kDa
Observed band size: 92 kDa why is the actual band size different from the predicted?

This data was developed using the same antibody clone in a different buffer formulation (ab68153).

  Lanes 1- 2: Merged signal (red and green). Green - ab68153 observed at 92 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

 ab68153 was shown to react with STAT3 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255436 (knockout cell lysate ab263797) was used. Wild-type HeLa and STAT3 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab68153 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human brain tissue sections labelling STAT3 with purified ab68153 at 1/200. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. A prediluted HRP-polymer conjugated anti-rabbit IgG was used as the secondary antibody. Counterstained with Hematoxylin.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).

Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling STAT3 (green) with unpurified ab68153 at 1/140. Cells were fixed with 4% paraformaldehyde. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/200) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).

Immunocytochemistry/Immunofluorescence analysis of HeLa cells labelling STAT3 (green) with purified ab68153 at 1/200. Cells were fixed with 4% paraformaldehyde. An Alexa Fluor^® 488-conjugated goat anti-rabbit IgG (1/200) was used as the secondary antibody. DAPI (blue) was used as the nuclear counterstain.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).

Flow cytometry analysis of Raji cells labelling STAT3 with unpurified ab68153 at 1/30 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. A rabbit monoclonal IgG was used as the isotype control (green).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).

Flow cytometry analysis of Raji cells labelling STAT3 with purified ab68153 at 1/50 (red). Cells were fixed with 2% paraformaldehyde. A FITC-conjugated goat anti-rabbit IgG (1/150) was used as the secondary antibody. A rabbit monoclonal IgG was used as the isotype control (green).

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab68153).