All lanes : Anti-RIP2 antibody [AF28D3] (ab75257) at 1/2000 dilution

Lane 1 : Wild-type HeLa cell lysate
Lane 2 : RIPK2 knockout HeLa cell lysate
Lane 3 : Ramos cell lysate
Lane 4 : Jurkat cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) at 1/20000 dilution

Predicted band size: 61 kDa
Observed band size: 65 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab75257 observed at 65 kDa. Red - loading control ab181602 observed at 37 kDa.

 ab75257 Anti-RIP2 antibody [AF28D3] was shown to specifically react with RIP2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab264688 (knockout cell lysate ab258636) was used. Wild-type and RIP2 knockout samples were subjected to SDS-PAGE. ab75257 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4°C at 1 in 2000 dilution and 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.

 

 

Allele-1: 2 bp deletion in exon 1.

 

Allele-2: 1 bp insertion in exon 1.