ab76009, at 1/100 dilution, staining B MyB in human urinary bladder carcinoma by immunohistochemistry using paraffin-embedded tissue.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

All lanes : Anti-B MyB (phospho T487) antibody [EPR2204Y] (ab76009) at 1/1000 dilution

Lane 1 : Raji cell lysate, untreated
Lane 2 : Raji cell lysate treated with TPA + IGF

Lysates/proteins at 10 µg per lane.

Secondary
All lanes : Goat anti-rabbit HRP at 1/1000 dilution

Predicted band size: 79 kDa
Observed band size: 110 kDa why is the actual band size different from the predicted?

ab76009, at 1/100 dilution, staining B MyB in HeLa cells by immunofluorescence.

Overlay histogram showing HeLa cells stained with ab76009 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab76009, 1/50 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monclonal) (1µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in HeLa cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.