All lanes : Anti-Importin4/Imp4 antibody [EPR13660] (ab181046) at 1/1000 dilution

Lane 1 : Wild-type HEK293T cell lysate
Lane 2 : IPO4 knockout HEK293T cell lysate
Lane 3 : HepG2 cell lysate
Lane 4 : A549 cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution

Predicted band size: 119 kDa
Observed band size: 130 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab181046 observed at 130 kDa. Red - loading control ab8245 observed at 36 kDa. 

 ab181046 Anti-Importin4/Imp4 antibody [EPR13660] was shown to specifically react with Importin4/Imp4 in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266175 (knockout cell lysate ab258005) was used.  Wild-type and Importin4/Imp4 knockout samples were subjected to SDS-PAGE.  ab181046 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively.  Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Allele-1: 10 bp deletion in exon4

 

Allele-2: 1 bp insertion in exon 4.