Anti-RNF31/HOIP antibody

• IP

Supplier Data

Immunoprecipitation - Anti-RNF31/HOIP antibody (AB125189)

Detection of human RNF31/HOIP by western blot of immunoprecipitates. Samples : Whole cell lysate (0.5 or 1.0 mg per IP reaction; 20% of IP loaded) from HeLa cells prepared using NETN lysis buffer. Antibodies : Affinity purified rabbit anti-RNF31/HOIP antibody ab125189 used for IP at 6 μg per reaction. RNF31/HOIP was also immunoprecipitated by a previous lot of this antibody. For blotting immunoprecipitated RNF31/HOIP, ab125189 was used at 1 μg/mL. Detection : Chemiluminescence with an exposure time of 3 minutes.

All lanes:

Immunoprecipitation - Anti-RNF31/HOIP antibody (ab125189)

Predicted band size: 120 kDa

false

• WB

Lab

Western blot - Anti-RNF31/HOIP antibody (AB125189)

Western blot : Anti-RNF31 antibody (ab125189) staining at 1/3000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab125189 was shown to bind specifically to RNF31. A band was observed at 130-140 kDa in wild-type A549 cell lysates with no signal observed at this size in RNF31 knockout cell line. To generate this image, wild-type and RNF31 knockout A549 cell lysates were analysed. First, 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-RNF31/HOIP antibody (ab125189) at 1/3000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

RNF31 knockout A549 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

HepG2 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

false

• WB

Lab

Western blot - Anti-RNF31/HOIP antibody (AB125189)

Western blot : Anti-RNF31 antibody (ab125189) staining at 1/1000 dilution, shown in green; Mouse anti-CANX [CANX/1543] (ab238078) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab125189 was shown to bind specifically to RNF31. A band was observed at 130 kDa in wild-type A549 cell lysates with no signal observed at this size in RNF31 knockout cell line. To generate this image, wild-type and RNF31 knockout A549 cell lysates were analysed. First, 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-RNF31/HOIP antibody (ab125189) at 1/1000 dilution

Lane 1:

Wild-type A549 cell lysate at 20 µg

Lane 2:

RNF31 knockout A549 cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

HepG2 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

Predicted band size: 120 kDa

Observed band size: 130 kDa

false

• WB

Unknown

Western blot - Anti-RNF31/HOIP antibody (AB125189)

All lanes:

Western blot - Anti-RNF31/HOIP antibody (ab125189) at 0.4 µg/mL

Lane 1:

HeLa whole cell lysate at 50 µg

Lane 2:

HeLa whole cell lysate at 15 µg

Lane 3:

293T whole cell lysate at 50 µg

Lane 4:

Jurkat whole cell lysate at 50 µg

Predicted band size: 120 kDa

true

Exposure time: 3min