All lanes : Anti-Beta Arrestin 2 antibody (ab54790) at 1 µg/ml

Lane 1 : Wild-type HepG2 cell lysate
Lane 2 : ARRB2 knockout HepG2 cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : Human Lung tissue lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 46 kDa
Observed band size: 50 kDa why is the actual band size different from the predicted?

Lanes 1 - 4: Merged signal (red and green). Green - ab54790 observed at 50 kDa. Red - loading control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) observed at 37 kDa.

ab54790 was shown to react with Beta Arrestin 2 in wild-type HepG2 cells in Western blot with loss of signal observed in ARRB2 knockout cell line ab262320 (knockout cell lysate ab257283). Wild-type HepG2 and ARRB2 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with ab54790 and ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) overnight at 4 °C at 1 µg/ml and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye^® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye^® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.

All lanes : Anti-Beta Arrestin 2 antibody (ab54790) at 1/500 dilution

Lane 1 : Wild-type HepG2 cell lysate
Lane 2 : ARRB2 knockout HepG2 cell lysate
Lane 3 : A549 cell lysate
Lane 4 : K562 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 46 kDa
Observed band size: 55 kDa why is the actual band size different from the predicted?

Lanes 1- 4: Merged signal (red and green). Green - ab54790 observed at 55 kDa. Red - Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) observed at 37 kDa.

 ab54790 was shown to react with ARRB2 in wild-type HepG2 cells in western blot. Loss of signal was observed when knockout cell line ab262320 (knockout cell lysate ab257283) was used. Wild-type HepG2 and ARRB2 knockout HepG2 cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab54790 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) overnight at 4°C at a 1 in 500 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Mouse IgG H&L (IRDye^®800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye^®680RD) preadsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Homozygous: 1 bp deletion in exon4