Anti-LIMK2 antibody [EP969Y] - C-terminal

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-LIMK2 antibody [EP969Y] - C-terminal (AB45165)

Immunocytochemistry/ Immunofluorescence analysis of Ramos cells labeling LIMK2 with ab45165 at 1/250. Goat anti rabbit IgG(Alexa Fluor® 488); ab150077 at 1/1000 dilution was used as the secondary antibody. Cells were fixed with 4% paraformaldehyde and permeabilised with 0.1% tritonX-100. DAPI (blue) was used as a nuclear counterstain.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LIMK2 antibody [EP969Y] - C-terminal (AB45165)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human endometrium cancer tissue labeling LIMK2 with ab45165 at 1/250 dilution. Heat mediated antigen retrieval was performed using Tris/EDTA buffer pH 9. ab97051, a HRP-conjugated goat anti-rabbit IgG (H+L) was used as the secondary antibody (1/500). Negative control using PBS instead of primary antibody. Counterstained with hematoxylin.

• WB

Supplier Data

Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (AB45165)

Lane 1 : Wild-type HAP1 whole cell lysate (40 μg)
Lane 2 : LIMK2 (LIMK2) knockout HAP1 whole cell lysate (40 μg)
Lane 3 : Jurkat whole cell lysate (40 μg)

Lanes 1 - 3 : Merged signal (red and green). Green - ab45165 observed at 72 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab45165 was shown to specifically recognize LIMK2 (LIMK2) in wild-type HAP1 cells along with additional cross-reactive bands. No band was observed with knockout cells were examined. Wild-type and LIMK2 (LIMK2) knockout samples were subjected to SDS-PAGE. ab45165 and ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution and 1/20,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed (ab216776) secondary antibodies at 1/20,000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (ab45165)

Predicted band size: 72 kDa

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• WB

Lab

Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (AB45165)

Western blot : Rabbit Monoclonal[EP969Y] to LIMK2 ab45165 staining at 1/2000 dilution, shown in green; Mouse anti GAPDH (ab8245) loading control staining at 1/20,000 dilution, shown in magenta.

A band was observed at 72 kDa in Wild-type U-87 MG cell lysates with no signal observed at this size in LIMK2 knockout U-87 MG cell line.

To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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 & Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (ab45165) at 1/1000 dilution

Lane 1:

Wild-type U-87 MG at 20 µg

Lane 2:

LIMK2 knockout U-87 MG cell line at 20 µg

Lane 3:

HCT 116 at 20 µg

Lane 4:

PC-3 at 20 µg

Secondary

All lanes:

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

Predicted band size: 72 kDa

Observed band size: 72 kDa

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• WB

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Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (AB45165)

All lanes:

Western blot - Anti-LIMK2 antibody [EP969Y] - C-terminal (ab45165) at 1/2000 dilution

All lanes:

Ramos (Human Burkitt's lymphoma cell line) membrane lysate

Predicted band size: 72 kDa

Observed band size: 70 kDa

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