Anti-Galectin 3 antibody [EPR2774]

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-Galectin 3 antibody [EPR2774] (AB76466)

ab76466, at a 1/100 dilution, staining Galectin 3 in HeLa cells by Immunofluorescence.

• Flow Cyt (Intra)

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Flow Cytometry (Intracellular) - Anti-Galectin 3 antibody [EPR2774] (AB76466)

Overlay histogram showing THP1 cells stained with ab76466 (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% human serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab76466 1/100 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor^® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1μg/1x10⁶ cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter.

• WB

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Western blot - Anti-Galectin 3 antibody [EPR2774] (AB76466)

All lanes:

Western blot - Anti-Galectin 3 antibody [EPR2774] (ab76466) at 1/5000 dilution

Lane 1:

HeLa cell lysate at 10 µg

Lane 2:

SW480 cell lysate at 10 µg

Lane 3:

A431 cell lysate at 10 µg

Lane 4:

MCF7 cell lysate at 10 µg

Secondary

All lanes:

goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 26 kDa

Observed band size: 30 kDa

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• WB

Lab

Western blot - Anti-Galectin 3 antibody [EPR2774] (AB76466)

Lane 1 : Wild type HAP1 whole cell lysate (20 μg)
Lane 2 : Galectin 3 (KO) knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HeLa whole cell lysate (20 μg)
Lane 4 : MCF7 whole cell lysate (20 μg)
Lanes 1 - 4 : Merged signal (red and green). Green - ab76466 observed at 35 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab76466 was shown to specifically react with Galectin 3 (KO) when Galectin 3 (KO) knockout samples were used. Wild-type and Galectin 3 (KO) knockout samples were subjected to SDS-PAGE. ab76466 and ab8245 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 2000 dilution and 1/10000 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 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Galectin 3 antibody [EPR2774] (ab76466)

Predicted band size: 26 kDa

Observed band size: 35 kDa

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