Anti-RBM15B 抗体 [EPR25177-134] (BSA and Azide free) (ab300468)

All lanes : Anti-RBM15B antibody [EPR25177-134] (ab300467) at 1/1000 dilution

Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) transfected with scrambled siRNA control, whole cell lysate
Lane 2 : HeLa transfected with siRNA specifically targeting RBM15B, whole cell lysate 'a'
Lane 3 : HeLa transfected with siRNA specifically targeting RBM15B, whole cell lysate 'b'

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: 97 kDa
Observed band size: 100 kDa why is the actual band size different from the predicted?


Exposure time: 114 seconds

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Blocking / Diluting buffer: 5% NFDM/TBST.

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID: 27602518).

The bands below 100 kDa may be caused by degradation.

All lanes : Anti-RBM15B antibody [EPR25177-134] (ab300467) at 1/1000 dilution

Lane 1 : His-tagged human RBM15B recombinant protein, 10 ng
Lane 2 : His-tagged human RBM15 recombinant protein, 10 ng

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Predicted band size: 97 kDa
Observed band size: 25 kDa why is the actual band size different from the predicted?

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Blocking / Diluting buffer: 5% NFDM/TBST.

This antibody does not cross-react with RBM15.

Both recombinant proteins were expressed from E.coli expression systems.

Exposure time: 7.75 seconds.

All lanes : Anti-RBM15B antibody [EPR25177-134] (ab300467) at 1/1000 dilution

Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : 293T (human embryonic kidney epithelial cell) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Predicted band size: 97 kDa
Observed band size: 100 kDa why is the actual band size different from the predicted?


Exposure time: 81 seconds

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Blocking / Diluting buffer: 5% NFDM/TBST.

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

The band below 100 kDa may be caused by degradation.

All lanes : Anti-RBM15B antibody [EPR25177-134] (ab300467) at 1/1000 dilution

Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate
Lane 2 : Raw264.7 (Mouse Abelson murine leukemia virus-induced tumor macrophage) whole cell lysate
Lane 3 : 2.4G2 (Rat B cell lymphoma B lymphocyte) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/50000 dilution

Developed using the ECL technique.

Predicted band size: 97 kDa
Observed band size: 100 kDa why is the actual band size different from the predicted?


Exposure time: 92 seconds

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Blocking and diluting buffer and concentration: 5% NFDM/TBST

Lysates were freshly made and used for Western blotting immediately to minimize protein degradation.

The band below 100 kDa may be caused by degradation.

This blot was developed using a higher sensitivity ECL substrate.

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded mouse cerebrum tissue labeling RBM15B with ab300467 at 1/100 dilution (4.92 µg/ml) followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. The section was incubated with ab300467 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Immunohistochemical analysis of paraffin-embedded mouse pancreatic carcinoma tissue labeling RBM15B with ab300467 at 1/100 dilution (4.92 µg/ml) followed by a ready to use Leica DS9800 (BOND™ Polymer Refine Detection) kit. The section was incubated with ab300467 for 30 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND^® RX instrument. Counterstained with Hematoxylin.

Secondary antibody only control: Secondary antibody is a ready to use LeicaDS9800 (BOND™ Polymer Refine Detection).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution2) for 20 mins.

This data was developed using ab300467, the same antibody clone in a different buffer formulation.

Flow Cytometry (Intracellular) analysis of HeLa (human cervix adenocarcinoma epithelial cells) labeling RBM15B (red) with ab300467 at a 1/50 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A Goat Anti-Rabbit IgG (Alexa Fluor^® 488, (ab150081)) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730) Isotype control. Blue - (unlabeled control) - Cells without incubation with primary and secondary antibodies.