Anti-PTEN antibody [Y184]

• WB

Lab

Western blot - Anti-PTEN antibody [Y184] (AB32199)

Lanes 1 : PTEN knockout HAP1 cell lysate (20 μg)
Lanes 2 : Wild-type HAP1 cell lysate (20 μg)
Green signal from target
Red signal from loading control – ab8245 observed at 37 kDa

This western blot image is a comparison between ab32199 and a competitor's top cited mouse monoclonal antibody.

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199)

Predicted band size: 47 kDa

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• WB

Unknown

Western blot - Anti-PTEN antibody [Y184] (AB32199)

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199) at 1/500 dilution

All lanes:

MCF7 cell lysate

Predicted band size: 47 kDa

Observed band size: 54 kDa

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• WB

Lab

Western blot - Anti-PTEN antibody [Y184] (AB32199)

Blocking buffer : 5% NFDM/TBST
Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199) at 1/10000 dilution

Lane 1:

MCF7 whole cell lysate at 10 µg

Lane 2:

HEK293 whole cell lysate at 10 µg

Secondary

All lanes:

Anti-rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 47 kDa

Observed band size: 54 kDa

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• WB

Lab

Western blot - Anti-PTEN antibody [Y184] (AB32199)

Lanes 1 and 5 : PTEN knockout HAP1 cell lysate (20 μg)
Lanes 2 and 6 : Wild-type HAP1 cell lysate (20 μg)
Lane 2 : Green signal from target – ab32199 observed at 47 kDa
Lanes 3 and 4 : Red signal from loading control – ab8245 observed at 37 kDa
Lanes 5 and 6 : Merged (red and green) signal

ab32199 was shown to specifically recognize PTEN in wild-type HAP1 cells along with additional cross reactive bands. No band was observed when PTEN knockout samples were used. Wild-type and PTEN knockout samples were subjected to SDS-PAGE, ab32199 and ab8245 (loading control to GAPDH) were diluted to 1/500 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 1 h at room temperature before imaging.

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199)

Predicted band size: 47 kDa

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• WB

AbReview43115****

Western blot - Anti-PTEN antibody [Y184] (AB32199)

Treatment :

Lane 1 - control siRNA 24 hours

Lane 2 - PTEN siRNA 24 hours

Lane 3 - control siRNA 48 hours

Lane 4 - PTEN siRNA 48 hours

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199) at 1/500 dilution

All lanes:

Mouse primary bone marrow derived macrophage whole cell lysate at 50 µg

Secondary

All lanes:

HRP-conjugated goat anti-rabbit IgG polyclonal

Predicted band size: 47 kDa

Observed band size: 50 kDa

true

Exposure time: 30s

This image is courtesy of an anonymous Abreview

• WB

AbReview50495****

Western blot - Anti-PTEN antibody [Y184] (AB32199)

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199) at 1/500 dilution

All lanes:

RBL-2H3 whole cell lysate

Secondary

All lanes:

Goat Anti-rabbit HRP at 1/1000 dilution

Predicted band size: 47 kDa

true

This image is courtesy of an anonymous Abreview.

• WB

Lab

Western blot - Anti-PTEN antibody [Y184] (AB32199)

Blocking buffer : 5% NFDM/TBST
Dilution buffer : 5% NFDM/TBST

All lanes:

Western blot - Anti-PTEN antibody [Y184] (ab32199) at 1/10000 dilution

All lanes:

Brain lysate at 10 µg

Secondary

All lanes:

Anti-rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/1000 dilution

Predicted band size: 47 kDa

Observed band size: 54 kDa

false

• WB

Lab

Western blot - Anti-PTEN antibody [Y184] (AB32199)

False colour image of Western blot : Anti-PTEN antibody [Y184] (ab32199) staining at 1/10000 dilution, shown in green; Mouse anti-GAPDH loading control staining, shown in magenta.

In Western blot, ab32199 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 [Y184] (ab32199) at 1/10000 dilution

Lane 1:

Wild-type VCAP cell lysates

Lane 2:

Western blot - Human PTEN knockout VCaP cell line (ab326417)

Secondary

All lanes:

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

Observed band size: 47 kDa

false