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ab8245 staining GAPDH in SV40LT-SMC (RatSV40-transfected aorta smooth cell line) cells.

The cells were fixed with 4% formaldehyde (10 minutes),permeabilized with 0.1% Triton X-100 for 5 minutes andthen blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for1 hour. The cells were then incubated with ab8245 at 5μg/ml and ab202272 at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1h withGoat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117)(shown in green). Nuclear DNA was labeled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab8245 staining GAPDH in NIH/3T3 (Mouse embryo fibroblast cell line) cells.

The cells were fixed with 4% formaldehyde (10 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 1 μg/ml and ab202272 at 1/250 overnight at +4°C, followed by a further incubation at room temperature for 1 hour withGoat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117)(shown in green). Nuclear DNA was labeled in blue with DAPI.

Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

ab8245 stainingGAPDH in HeLa (Human epithelial cell line from cervix adenocarcinoma) cells.

The cells were fixed with 100% methanol (5 minutes) and then blocked in 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1 hour. The cells were then incubated with ab8245 at 5 μg/ml and ab6046 at 1 µg/ml overnight at +4°C, followed by a further incubation at room temperature for 1 hour withGoat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117)at 2 μg/ml (shown in green) andGoat Anti-Rabbit IgG H&L (Alexa Fluor^® 594) preadsorbed (ab150088)at 2 μg/ml (shown in pseudo color red). Nuclear DNA was labeled in blue with DAPI.

Negative controls: 1– Rabbit primary antibodyand anti-mouse secondary antibody; 2 – Mouse primary antibody and anti-rabbit secondary antibody. Controls 1 and 2 indicate that there is no unspecific reaction between primary and secondary antibodies used.