All lanes : Anti-FANCI antibody (ab103077) at 1/500 dilution

Lane 1 : Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 2 : FANCI knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 4 : Raji (Human Burkitts lymphoma cell line) whole 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: 149 kDa
Observed band size: 151 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab103077 observed at 151 kDa. Red - loading control ab8245 observed at 36 kDa.

 ab103077 Anti-FANCI antibody was shown to specifically react with FANCI in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266278 (knockout cell lysate ab257434) was used. Wild-type and FANCI knockout samples were subjected to SDS-PAGE. ab103077 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 500 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: 2 bp deletion in exon 4

 

Allele-2: Insertion of the selection cassette in exon 4.