Recombinant Anti-WFS1 antibody [EPR23801-91] (ab259362)

IHC image of WFS1 staining in a section of frozen normal human pancreas* performed on a Leica BOND™ system using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab259362, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

IHC image of WFS1 staining in a section of frozen normal human hippocampus* performed on a Leica BOND™ system using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab259362, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

Immunohistochemical analysis of paraffin-embedded Mouse pancreas tissue labelling WFS1 with ab259362 at 1/5000 dilution (0.093 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on mouse pancreatic islet (PMID: 15994758). The section was incubated with ab259362 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 Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HepG2 cells labelling WFS1 with ab259362 at 1/50 dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) antibody at 1/1000 dilution (Green). Confocal image showing cytoplasmic staining in HepG2 cells is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) at 1/1000 dilution.

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse pancreas tissue labeling WFS1 with ab259362 at 1/50 (9.28 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (Green). Positive staining on mouse pancreatic islets is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488)at 1/1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

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

The observed MW is consistent with what has been described in the literature (PMID: 12913071). 

Lysates were made freshly and used in WB test immediately to minimize protein degradation.

Samples (except lane1) are non-boiled as boiling may cause protein aggregates.

Exposure time: Lane 1: 10 secondsLane 2-5: 3 minutes

WFS1 was immunoprecipitated from 0.35 mg Mouse eyeball tissue lysate with ab259362 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab259362 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.

Lane 1: Mouse eyeball tissue lysate 40 ug

Lane 2: ab259362 IP in Mouse eyeball tissue lysate

Lane 3:Rabbit monoclonal IgG (ab172730) instead of ab259362 in mouse eyeball tissue lysate

Blocking and dilution buffer and concentration: 5% NFDM/TBST.

Exposure time: 50 seconds

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

The observed MW is consistent with what has been described in the literature (PMID: 12913071 ).

Samples are non-boiled as boiling may cause protein aggregates. 

 Exposure time: 8 seconds

Immunohistochemical analysis of paraffin-embedded Mouse cerebrum tissue labelling WFS1 with ab259362 at 1/5000 dilution (0.093 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on the CA1 region of mouse hippocampus (PMID: 24694561). The section was incubated with ab259362 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 Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

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

The observed MW is consistent with what has been described in the literature (PMID: 12913071).

Low expression: spleen, PC-12 (PMID: 12913071; PMID: 11181571 ).

Samples are non-boiled as boiling may cause protein aggregates.

Exposure time: 8 seconds

Immunohistochemical analysis of paraffin-embedded Rat pancreas tissue labelling WFS1 with ab259362 at 1/5000 dilution (0.093 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on rat pancreatic islet. The section was incubated with ab259362 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 Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen rat pancreas tissue labeling WFS1 with ab259362 at 1/50 (4.64 ug/ml) dilution followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution (Green). Positive staining on rat pancreatic islets is observed. The nuclear counterstain was DAPI (Blue).

Secondary antibody control: Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488)at 1/1000 dilution.

Heat mediated antigen retrieval using sodium citrate buffer (10mM citrate pH 6.0 + 0.05% Tween-20).

Immunohistochemical analysis of paraffin-embedded Rat cerebrum tissue labelling WFS1 with ab259362 at 1/5000 dilution (0.093 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Cytoplasmic staining on the CA1 region of rat hippocampus. The section was incubated with ab259362 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 Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins

Immunohistochemical analysis of paraffin-embedded Mouse spleen tissue labelling WFS1 with ab259362 at 1/5000 dilution (0.093 ug/ml) followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). Negative control: almost no staining on mouse spleen. The section was incubated with ab259362 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 Citrate buffer (pH 6.0, epitope retrieval solution 1) for 20 mins