Anti-Von Hippel Lindau/VHL antibody [EPR30583-645]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of paraffin-embedded Human testis tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/100 (5.22 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human testis. The primary antibody was incubated 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 was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of paraffin-embedded Human colon carcinoma tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/100 (5.22 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human colon carcinoma. The primary antibody was incubated 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 was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of paraffin-embedded Human breast carcinoma tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/100 (5.22 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on human breast carcinoma. The primary antibody was incubated 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 was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-Fr

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Immunohistochemistry (Frozen sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of 4% PFA-fixed, 0.2% Triton X-100 permeabilized frozen Mouse colon (perfused fixed) tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/50 (10.44 ug/ml) dilution followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed at 1/1000 2 ug/mL dilution (Green).

Confocal image showing positive staining on mouse colon. The nuclear counterstain was DAPI (Blue). The section was incubated with the primary antibody for 60 mins at room temperature. The section was then mounted using Fluoromount®.The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

Secondary antibody control : Secondary antibody is ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbedat 1/1000 2 ug/mL dilution.

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

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of paraffin-embedded Mouse colon tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/100 (5.22 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse colon. The primary antibody was incubated 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 was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

Immunohistochemical analysis of paraffin-embedded Mouse breast carcinoma tissue labeling Von Hippel Lindau/VHL with ab324676 at 1/100 (5.22 ug/ml) dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection).

Positive staining on mouse breast carcinoma. The primary antibody was incubated 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 was performed with Tris-EDTA buffer (pH 9.0, Epitope Retrieval Solution2) for 20 mins

• WB

Supplier Data

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

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

Low expression : 786-O(PMID : 20300531)

All lanes:

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (ab324676) at 1/1000 dilution

Lane 1:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Lane 2:

786-O (human kidney epithelial cell) whole cell lysate at 40 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 18 kDa,22 kDa,124 kDa

false

Exposure time: 114s

• WB

Supplier Data

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

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

The expression of VHL is upregulated in response to VH298 treatment (PMID : 27811928).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Exposure time : Lane1-2 : 15 seconds, lane 3-5 : 136 seconds

All lanes:

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (ab324676) at 1/1000 dilution

Lane 1:

Untreated HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 2:

HeLa treated with 150 μM VH298 for 24 hours whole cell lysate at 20 µg

Lane 3:

Human colon tissue lysate at 20 µg

Lane 4:

Human cerebellum tissue lysate at 20 µg

Lane 5:

Human testis tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 18 kDa,22 kDa,36 kDa

false

• WB

Supplier Data

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

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

This antibody does not cross-react with recombinant human VHLL by western blot.

In Western blot, Anti-Myc tag® antibody staining at 1/5000 dilution.

All lanes:

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (ab324676) at 1/1000 dilution

Lane 1:

293T (human embryonic kidney epithelial cell) cells transfected with an empty vector containing a Myc-his tag, whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human VHL expression vector containing a Myc-his tag, whole cell lysate at 10 µg

Lane 3:

293T cells transfected with a human VHLL expression vector containing a Myc-his tag, whole cell lysate at 40 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 18-36 kDa,36 kDa

false

Exposure time: 10s

• WB

Supplier Data

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

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

The expression of VHL is upregulated in response to VH298 treatment (PMID : 27811928).

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 15899860)

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Exposure time : Lane1-2 : 180 seconds, lane 3 : 15 seconds

All lanes:

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (ab324676) at 1/1000 dilution

Lane 1:

Untreated NIH/3T3 (mouse embryonic fibroblast) whole cell lysate at 20 µg

Lane 2:

NIH/3T3 treated with 150 μM VH298 for 24 hours whole cell lysate at 20 µg

Lane 3:

Mouse testis tissue lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 19 kDa,20 kDa,36 kDa

false

• WB

Supplier Data

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (AB324676)

In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution.

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

Performed under reducing conditions. ab324676 was shown to bind specifically to VHL. Target of interest was observed at 19, 20 kDa in wild-type Renca cell lysates (lane 1) with no signal observed at this size in VHL knockout cell line (lane 2).

All lanes:

Western blot - Anti-Von Hippel Lindau/VHL antibody [EPR30583-645] (ab324676) at 1/1000 dilution

Lane 1:

Renca (mouse kidney epithelial) whole cell lysate at 50 µg

Lane 2:

VHL KO Renca whole cell lysate at 50 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 19-20 kDa,36 kDa

false

Exposure time: 180s