Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human kidney tissue sections labeling IL-11RA with purified ab125015 at 1:250 dilution (3.2 µg/ml). Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Goat Anti-Rabbit IgG H&L (HRP) ab97051
was used as the secondary antibody. Negative control:PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

Immunocytochemistry/ Immunofluorescence analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labeling IL-11RA with Purified ab125015 at 1:100 dilution (8.1 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.

All lanes : Anti-IL-11RA antibody [EPR5446] (ab125015) at 1/1000 dilution

Lane 1 : RAW264.7 (mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 2 : WEHI-231 (mouse B cell lymphoma B lymphocyte) whole cell lysate
Lane 3 : M1 (mouse myeloid leukemia myeloblast) whole cell lysate
Lane 4 : Mouse spleen tissue lysate
Lane 5 : Mouse testis tissue lysate
Lane 6 : Mouse thymus tissue lysate
Lane 7 : Mouse lymph node tissue lysate

Lysates/proteins at 20 µg per lane.

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

Predicted band size: 45 kDa
Observed band size: 42,45 kDa why is the actual band size different from the predicted?


Exposure time: 3 minutes

The expression pattern is consistent with what has been described in the literature (PMID: 26876177).

Blocking Buffer and concentration: 5% NFDM/TBST

All lanes : Anti-IL-11RA antibody [EPR5446] (ab125015) at 1/10000 dilution (Purified)

Lane 1 : A673 (Human muscle Ewing's Sarcoma ) whole cell lysates
Lane 2 : PC-3 (Human prostate adenocarcinoma epithelial cell) whole cell lysates
Lane 3 : HuT-78 (Human Sezary syndrome cutaneous T lymphocyte) whole cell lysates
Lane 4 : K-562 (Human chronic myelogenous leukemia lymphoblast) whole cell lysates

Lysates/proteins at 20 µg per lane.

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

Predicted band size: 45 kDa
Observed band size: 45 kDa

Flow Cytometry analysis of K-562 (Human chronic myelogenous leukemia lymphoblast) cells labeling IL-11RA with Purified ab125015 at 1:80 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).

ELISA analysis of Human IL-11RA recombinant protein at 1000 ng/mL with ab125015. An Alkaline Phosphatase-conjugated AffiniPure Goat Anti-Rabbit IgG (H+L) at 1/2500 dilution was used as the secondary antibody.

Overlay histogram showing K562 cells stained with unpurified ab125015 (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 (ab125015, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. 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 K562 cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

All lanes : Anti-IL-11RA antibody [EPR5446] (ab125015) at 1/1000 dilution (Unpurified)

Lane 1 : 293T lysate
Lane 2 : Saos2 lysate
Lane 3 : A673 lysate
Lane 4 : PC3 lysate
Lane 5 : Hut78 lysate
Lane 6 : K562 lysate
Lane 7 : TF1 lysate

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat anti-Rabbit HRP at 1/2000 dilution

Predicted band size: 45 kDa
Observed band size: 45 kDa

Unpurified ab125015 at 1/500 staining IL-11RA in paraffin embedded Human Kidney tissue by immunohistochemistry.

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

Equilibrium disassociation constant (K_D)
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