重组Anti-PTEN抗体[EPR9941]

• WB

Lab

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

Western blot : Anti-PTEN antibody [EPR9941] ab154812 staining at 1/10000 dilution, shown in green; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 50 kDa in Wild-type A549 and HeLa cell lysates with no signal observed at this size in PTEN knockout A549 cell line and PTEN knockout HeLa cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 5 % Milk in TBS-0.1 % Tween^® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/10000 dilution

Lane 1:

Wild-type A549 cell lysates at 20 µg

Lane 2:

Western blot - Human PTEN knockout A549 cell line (<a href='/products/cell-lines/human-pten-knockout-a549-cell-line-ab286704'>ab286704</a>) at 20 µg

Lane 3:

Wild-type HeLa ab255928 cell lysates at 20 µg

Lane 4:

Western blot - Human PTEN knockout HeLa cell line (<a href='/products/cell-lines/human-pten-knockout-hela-cell-line-ab255419'>ab255419</a>) at 20 µg

Lane 5:

MCF7 cell lysates at 20 µg

Lane 6:

LNCaP cell lysates at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 47 kDa

Observed band size: 50 kDa

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• WB

Lab

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

False colour image of Western blot : Anti-PTEN antibody [EPR9941] (ab154812) staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH loading control staining, shown in magenta.

In Western blot, ab154812 was shown to bind specifically to PTEN. A band was observed at 47 kDa in Wild-type VCAP cell lysates with no signal observed at this size in PTEN knockout cell line.

Samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3% milk in TBS-0.1% Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°C.

Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged.

Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed at 1 : 10000 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed at 1 : 10000.

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/10000 dilution

Lane 1:

Wild-type VCAP cell lysates

Lane 2:

Western blot - Human PTEN knockout VCaP cell line (<a href='/products/cell-lines/human-pten-knockout-vcap-cell-line-ab326417'>ab326417</a>)

Secondary

All lanes:

Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed at 1/10000 dilution

Observed band size: 47 kDa

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• WB

Unknown

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/1000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

MCF7 cell lysate at 10 µg

Lane 3:

293T cell lysate at 10 µg

Lane 4:

A431 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 47 kDa

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• WB

Lab

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

Lanes 1- 2 : Merged signal (red and green). Green - ab154812 observed at 47 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

ab154812 was shown to react with PTEN in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255419 (knockout cell lysate ab263829) was used. Wild-type HeLa and PTEN knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab154812 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 10000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/10000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

PTEN knockout HeLa cell lysate at 20 µg

Predicted band size: 47 kDa

Observed band size: 47 kDa

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• WB

Lab

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

Lanes 1 and 2 : Merged signal (red and green). Green - ab154812 observed at 51 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab154812 was shown to specifically recognize PTEN in wild-type HAP1 cells. No band was observed when PTEN knockout samples were used. Wild-type and PTEN knockout samples were subjected to SDS-PAGE, ab154812 and ab8245 (loading control to GAPDH) were diluted 1/1000 and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1/10,000 dilution for 1hr at room temperature before imaging.

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/1000 dilution

Lane 1:

Wild-type HAP1 cell lysate at 20 µg

Lane 2:

PTEN knockout HAP1 cell lysate at 20 µg

Predicted band size: 47 kDa

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• WB

Supplier Data

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/1000 dilution

Lane 1:

Mouse brain Lysate at 10 µg

Lane 2:

Rat brain Lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), HRP- conjugated at 1/1000 dilution

Predicted band size: 47 kDa

Observed band size: 54 kDa

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• WB

Supplier Data

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

All lanes:

Western blot - Anti-PTEN antibody [EPR9941] (ab154812) at 1/1000 dilution

Lane 1:

HeLa cell Lysate at 10 µg

Lane 2:

Neuro-2a cell Lysate at 10 µg

Lane 3:

Rat spleen cell Lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), HRP- conjugated at 1/1000 dilution

Predicted band size: 47 kDa

Observed band size: 54 kDa

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• WB

CiteAb

Western blot - Anti-PTEN antibody [EPR9941] (AB154812)

PTEN western blot using anti-PTEN antibody [EPR9941] ab154812. Publication image and figure legend from Noll, A., Illuzzi, G., et al., 2016, Cancer Cell Int, PubMed 27375368.

ab154812 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab154812 please see the product overview.

PARG deficiency is not synthetic lethal with PTEN deficiency. a The PTEN-mutated MDA-MB-468 cell line is not sensitive to PARG depletion. Clonogenic survival (left panel) of MDA-MB-468 cells after siCTL and siPARG transfection. Results are depicted as box plots showing distribution of data from 5 individual experiments. Relative cell number 72 h post-siRNA transfection (right panel) is counted from 5 individual experiments. PARG depletion was verified by western blot (middle panel). b Clonogenic survival (left panel) of MDA-MB-231 cells after single or combined siRNA-mediated depletion of BRCA1, PARG and PTEN. Results are depicted as box plots showing distribution of data from 5 independent experiments. Percentage of viable cells relative to siCTL-transfected cells 72 h post-transfection (right panel) is counted from 5 individual experiments. BRCA1, PARG and PTEN siRNA-mediated depletions compared to untransfected or siCTL transfected cells were verified by western blot (middle panel). The arrow points to BRCA1 signal above a non-specific band (#). p : 0.01 < * < 0.05; 0.05 < ns, not significant

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