Anti-Talin antibody [8D4]

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Talin antibody [8D4] (AB157808)

Overlay histogram showing RAW 264.7 cells stained with ab157808 (red line). The cells were fixed with 80% methanol (5 min)/ and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab157808, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in RAW 264.7 cells fixed with 4% formaldehyde (10 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.

• WB

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Western blot - Anti-Talin antibody [8D4] (AB157808)

Western blot membranes were incubated with diluted antibody in 5% non-fat milk, PBS and 0.04% Tween20 for 1 hour at room temperature.

Lanes 1 - 2:

Western blot - Anti-Talin antibody [8D4] (ab157808) at 1/1000 dilution

Lanes 3 - 4:

Anti-Talin 1 (phospho S425) antibody (<a href='/en-us/products/unavailable/talin-1-phospho-s425-antibody-ab157807'>ab157807</a>) at 1/1000 dilution

Lanes 1 and 3:

PC12 cell lysate - stimulated with calyculin A at 100nM for 30 minutes

Lanes 2 and 4:

PC12 cell lysate - stimulated with calyculin A at 100nM for 30 minutes and treated with lambda phosphatase

Predicted band size: 269 kDa

Observed band size: 240 kDa

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• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Talin antibody [8D4] (AB157808)

Immunocytochemical analysis of chick fibroblasts labelling Talin 1 with ab157808 at 1/100 (red). F-actin was stained in green.
Image provided by Dr. Gianluca Gallo at Drexel University.

• WB

Lab

Western blot - Anti-Talin antibody [8D4] (AB157808)

Anti-Talin antibody [8D4] ab157808 staining at 1/1000 dilution, shown in green; Rabbit anti-ACTN4 (ab108198) loading control staining at 1/20,000 dilution, shown in magenta. Bands were observed at 220 and 240 kDa. 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-Mouse 800CW and Goat anti-Rabbit 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Talin antibody [8D4] (ab157808) at 1/1000 dilution

Lane 1:

PC-3 cell lysate at 20 µg

Lane 2:

HeLa cell lysate at 20 µg

Lane 3:

SK-OV-3 cell lysate at 20 µg

Secondary

All lanes:

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

Predicted band size: 269 kDa

Observed band size: 220 kDa,240 kDa

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