Recombinant Anti-SATB1 antibody [EPR3951] (ab109122)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human lung carcinoma tissue sections labeling SATB1 with Purified ab109122 at 1:500 dilution (0.246 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of mouse liver tissue sections labeling SATB1 with Purified ab109122 at 1:500 dilution (0.246 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of rat liver tissue sections labeling SATB1 with Purified ab109122 at 1:500 dilution (0.246 µg/ml). Heat mediated antigen retrieval was performed using Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Tissue was counterstained with Hematoxylin. Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) secondary antibody was used at 1:0 dilution. PBS instead of the primary antibody was used as the negative control. 

Purified ab109122 at 1:20 dilution (0.6µg) immunoprecipitating SATB1 in Rat thymus lysate.
Lane 1 (input): Rat thymus lysate 10 µg
Lane 2 (+): ab109122 + Rat thymus lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab109122 in Rat thymus lysate.
VeriBlot for IP Detection Reagent (HRP)(ab131366) (1:5000 dilution) was used for Western blotting.
Blocking Buffer and concentration: 5% NFDM/TBST.
Diluting buffer and concentration: 5% NFDM/TBST.
Observed band size: kDa

Flow Cytometry analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labelling SATB1 with Purified ab109122 at 1:20 dilution (10 µg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488, ab150077) secondary antibody was used at 1:2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabelled control - Cell without incubation with primary antibody and secondary antibody (Blue).

Immunocytochemistry analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling SATB1 with Purified ab109122 at 1:50 dilution (2.5 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 (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: SATB1 knockout HAP1 whole cell lysate (20 µg)
Lane 3: Jurkat whole cell lysate (20 µg)
Lane 4: THP1 whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab109122 observed at 100 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab109122 was shown to specifically react with SATB1 in wild-type HAP1 cells as signal was lost in SATB1 knockout cells. Wild-type and SATB1 knockout samples were subjected to SDS-PAGE. Ab109122 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/1000 dilution and 1/20000 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 1/20000 dilution for 1 hour at room temperature before imaging.