All lanes : Anti-Hsp90 antibody [D7a] (ab59459) at 1/1000 dilution

Lane 1 : Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 2 : HSP90AA1 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 : HAP1 whole cell lyate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) at 1/10000 dilution

Predicted band size: 85 kDa
Observed band size: 90 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab59459 observed at 90 kDa. Red - loading control ab181602 observed at 36 kDa.

 ab59459 Anti-Hsp90 antibody [D7a] was shown to specifically react with Hsp90 in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266591 (knockout cell lysate ab258458) was used. Wild-type and Hsp90 knockout samples were subjected to SDS-PAGE. ab59459 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) 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^® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye^® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes : Anti-Hsp90 alpha antibody [2G5.G3] (ab79849) at 1/1000 dilution

Lane 1 : Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
Lane 2 : HSP90AA1 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 : HAP1 whole cell lyate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) at 1/10000 dilution

Predicted band size: 85 kDa
Observed band size: 90 kDa why is the actual band size different from the predicted?

Lanes 1-4: Merged signal (red and green). Green - ab79849 observed at 90 kDa. Red - loading control ab181602 observed at 36 kDa.

 ab79849 Anti-Hsp90 alpha antibody [2G5.G3]  was shown to specifically react with Hsp90 alpha in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266591 (knockout cell lysate ab258458) was used. Wild-type and Hsp90 alpha knockout samples were subjected to SDS-PAGE. ab79849 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) 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^® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye^® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

Homozygous: 1 bp insertion in exon3