Lane 1: Wild-type HAP1 whole cell lysate (20 µg)
Lane 2: FKBP51 knockout HAP1 whole cell lysate (20 µg)
Lane 3: HeLa whole cell lysate (20 µg)
Lane 4: HepG2 whole cell lysate (20 µg)

Lanes 1 - 4: Merged signal (red and green). Green - ab126715 observed at 51 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab126715 was shown to recognize FKBP51 in wild-type cells as signal was lost at the expected MW in FKBP51 knockout cells. Additional cross-reactive bands were observed in the wild-type and knockout cells. Wild-type and FKBP51 knockout samples were subjected to SDS-PAGE. Ab126715 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.

All lanes : Anti-FKBP51 antibody [EPR6617] (ab126715) at 1/10000 dilution (Purified)

Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysates at 15 µg
Lane 2 : Rat spleen lysates at 15 µg


Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution

Predicted band size: 51 kDa
Observed band size: 51 kDa

ab126715 (purified) at 1:20 dilution (2µg) immunoprecipitating FKBP51 in Jurkat whole cell lysate.
Lane 1 (input): Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10µg
Lane 2 (+): ab126715 & Jurkat whole cell lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab126715 in Jurkat 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.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Rat stomach tissue sections labeling FKBP51 with purified ab126715 at 1:250 dilution (1.156 µg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control:PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human prostatic hyperplasia tissue sections labeling FKBP51 with purified ab126715 at 1:250 dilution (1.156 µg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use)was used as the secondary antibody. Negative control:PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

Immunocytochemistry/ Immunofluorescence analysis of Jurkat (Human T cell leukemia T lymphocyte) cells labeling FKBP51 with purified ab126715 at 1:100 dilution (2.9 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.

Alexa Fluor^® 488 (ab198978) and Alexa Fluor^® 647 (ab198979) conjugated versions are available for this clone.

Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling FKBP51 with purified ab126715 at 1:20 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1:2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

Alexa Fluor^® 488 (ab198978) and Alexa Fluor^® 647 (ab198979) conjugated versions are available for this clone.

All lanes : Anti-FKBP51 antibody [EPR6617] (ab126715) at 1/1000 dilution

Lane 1 : Human testis cell lysate
Lane 2 : Jurkat cell lysate
Lane 3 : Caco-2 cell lysate
Lane 4 : HeLa cell lysate

Predicted band size: 51 kDa

ab126715, at 1/50 dilution, stains FKBP51 in paraffin embedded human colon tissue by immunohistochemistry.

Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

Equilibrium disassociation constant (K_D)
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