ab16802 at a 1/2000 dilution staining approx. 42kDa Glutamine synthetase in 1) 293T cell lysate transfected with Myc-Glutamine synthetase, 2) mouse brain lysate, 3) mouse liver lysate and 4) rat brain lysate by Western blot (ECL). ab16802 at a 1/2000 dilution staining approx. 42kDa Glutamine synthetase in 1) 293T cell lysate transfected with Myc-Glutamine synthetase, 2) mouse brain lysate, 3) mouse liver lysate and 4) rat brain lysate by Western blot (ECL).

Left panel shows scanned image of an amine coated plate seeded with a titration of HeLa cells and stained with different dilutions of ab16082 using In Cell ELISA support pack ab111542 and protocol. Right panel shows bar graph of normalized signal intensity in 3 different cells lines after staining with ab160802 at 1/2500 dilution. Average intra-assay coefficient of variation was calculated at 5.6% and Z score at 0.77.

HeLa cells were seeded on glass coverslips and allow to adhere overnight. Cells were fixed with 4% paraformaldehyde for 15 min and washed in PBS x3. ICC procedure was carried out using In Cell ELISA support pack ab111541 and protocol. Left panel shows HeLa cells stained with ab16802 at a dilution of 1/1000, labeled with Goat anti-rabbit 594 secondary antibody and counterstain with DAPI. Right panel shows co-localization of the signal with the mitochondrial marker Anti-ATPB antibody [3D5] ab14730 (4µg/mL) labeled with Goat anti-mouse 488 secondary antibody.

ICC/IF image of ab16802 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 (ab16802, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit 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.