Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (AB325018)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized HUVEC (human umbilical vein endothelial cell) cells labelling VE Cadherin (phospho Y685) with ab325018 at 1/1000 (0.495 ug/ml) dilution, followed by ab150081 Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed antibody at 1/1000 (2 ug/ml) dilution (Green).

Confocal image showing membranous and cytoplasmic staining in HUVEC cells (shown in green) treated with 100 uM Pervanadate for 10 mins, and showing no staining in untreated HUVEC cells. The counterstain was observed in magenta. Nuclear DNA was labelled with DAPI (shown in blue). Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab7291 Anti-alpha Tubulin mouse monoclonal antibody was used to counterstain tubulin at 1/1000 (1 ug/ml) dilution, followed by ab150120 Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) at 1/1000 (2 ug/ml) dilution (Magenta).

• WB

Supplier Data

Western blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (AB325018)

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

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

In Western blot, Anti-VE Cadherin antibody [EPR27436-55] (ab313633) staining at 1/1000 dilution.

All lanes:

Western blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (ab325018) at 1/1000 dilution

Lane 1:

Untreated HUVEC (human umbilical vein endothelial cell) whole cell lysate (untreated membrane) at 20 µg

Lane 2:

HUVEC treated with 100 μM Pervanadate for 10 minutes whole cell lysate (untreated membrane) at 20 µg

Lane 3:

Untreated HUVEC whole cell lysate (alkaline phosphatase treated membrane) at 20 µg

Lane 4:

HUVEC treated with 100 μM Pervanadate for 10 minutes whole cell lysate (alkaline phosphatase treated membrane) 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: 150 kDa,36 kDa

false

Exposure time: 26s

• WB

Supplier Data

Western blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (AB325018)

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

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

In Western blot, Anti-6X His tag® antibody [EPR20547] - ChIP Grade (ab213204) staining at 1/5000 dilution.

All lanes:

Western blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (ab325018) at 1/5000 dilution

Lane 1:

293T cells transfected with an empty vector containing a his-tag, whole cell lysate at 5 µg

Lane 2:

293T cells transfected with a human wild-type VE Cadherin expression vector containing a his-tag were treated with 100uM Pervanadate for 10 mins, whole cell lysate at 5 µg

Lane 3:

293T cells transfected with a human VE Cadherin (Y685A mutant) expression vector containing a his-tag were treated with 100uM Pervanadate for 10 mins, whole cell lysate at 5 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: 150 kDa,36 kDa

false

Exposure time: 180s

• Dot

Supplier Data

Dot Blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (AB325018)

Dot blot analysis of VE Cadherin (phospho Y685) using ab325018 at 1 : 1000 (0.495 ug/ml) followed by a Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (ab97051) at 1 : 100,000 dilution.

Lane 1 : VE Cadherin (phospho Y685) peptide
Lane 2 : VE Cadherin non-phospho peptide

Exposure time : 81 seconds.

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

All lanes:

Dot Blot - Anti-VE Cadherin (phospho Y685) antibody [EPR27437-539] (ab325018) at 1/1000 dilution

Lane 1:

VE Cadherin (phospho Y685) peptide

Lane 2:

VE Cadherin non-phospho peptide

Secondary

All lanes:

Dot 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

false

Exposure time: 81s