Anti-IMP3 antibody [EPR12022]

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-IMP3 antibody [EPR12022] (AB177942)

Immunofluorescent analysis of HeLa cells labeling IMP3 with ab177942 at 1/50 dilution (green). DAPI nuclear staining (blue).

• IP

Supplier Data

Immunoprecipitation - Anti-IMP3 antibody [EPR12022] (AB177942)

Western blot analysis on immunoprecipitation pellet from 293T cell lysate using ab177942.

All lanes:

Immunoprecipitation - Anti-IMP3 antibody [EPR12022] (ab177942)

Predicted band size: 63 kDa

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• WB

Lab

Western blot - Anti-IMP3 antibody [EPR12022] (AB177942)

Western blot : Anti-IGF2BP3 antibody [EPR12022] (ab177942) staining at 1/2000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab177942 was shown to bind specifically to IGF2BP3. A band was observed at 60-75 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in IGF2BP3 knockout cell line. To generate this image, wild-type and IGF2BP3 knockout HCT 116 cell lysates were analysed. First, 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 IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-IMP3 antibody [EPR12022] (ab177942) at 1/2000 dilution

Lane 1:

Wild-type HCT 116 cell lysate at 20 µg

Lane 2:

IGF2BP3 knockout HCT 116 cell lysate at 20 µg

Lane 3:

Wild-type HAP1 cell lysate at 20 µg

Lane 4:

IGF2BP3 knockout HAP1 cell lysate at 20 µg

Secondary

All lanes:

Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

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• WB

Lab

Western blot - Anti-IMP3 antibody [EPR12022] (AB177942)

Lane 1 : Wild-type HAP1 whole cell lysate (20 μg)
Lane 2 : IGF2BP3 (IMP3) knockout HAP1 whole cell lysate (20 μg)
Lane 3 : HeLa whole cell lysate (20 μg)
Lane 4 : HEK293 whole cell lysate (20 μg)

Lanes 1 - 4 : Merged signal (red and green). Green - ab177942 observed at 70 kDa. Red - loading control, ab9484, observed at 37 kDa.

ab177942 was shown to specifically react with IGF2BP3 (IMP3) in wild-type HAP1 cells as signal was lost in IGF2BP3 (IMP3) knockout cells. Wild-type and IGF2BP3 (IMP3) knockout samples were subjected to SDS-PAGE. ab177942 and ab9484 (Mouse anti GAPDH loading control) were incubated overnight at 4°C at 1/1000 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-IMP3 antibody [EPR12022] (ab177942)

Predicted band size: 63 kDa

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• WB

Supplier Data

Western blot - Anti-IMP3 antibody [EPR12022] (AB177942)

All lanes:

Western blot - Anti-IMP3 antibody [EPR12022] (ab177942) at 1/1000 dilution

Lane 1:

Hela cell lysate at 10 µg

Lane 2:

293T cell lysate at 10 µg

Lane 3:

K562 cell lysate at 10 µg

Lane 4:

HT-29 cell lysate at 10 µg

Predicted band size: 63 kDa

Observed band size: 69 kDa

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