All lanes : Anti-G gamma4 antibody (ab21789) at 1/1000 dilutionLane 1 : Total protein extract from E. coli with ~50ng to 100ng of a negative control fusion protein with an irrelevant antigen at 20 ug Lane 2 : Total protein extract from E. coli with ~50ng to 500ng of the antigen fusion protein at 20 ugSecondaryRabbit anti-mouse IgG + IgM, (H+L) horseradish peroxidase conjugated at 1/5000 dilution
ICC/IF image of ab21789 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab21789, 1/1000 dilution) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.