Abcam is committed to meeting high standards of ethical manufacturing and has decided to discontinue this product by June 2019 as it has been generated by the ascites method. We are sorry for any inconvenience this may cause.
Our Abpromise guarantee covers the use of ab58301 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 1 - 5 µg/ml. Predicted molecular weight: 62 kDa.|
|IHC-P||Use a concentration of 3 µg/ml.|
|ICC/IF||Use a concentration of 1 µg/ml.|
Lanes 1, 3 and 5: PKR knockout HAP1 cell lysate (20 µg)
Lanes 2, 4 and 6: Wild-type HAP1 cell lysate (20 µg)
Lanes 1 and 2: Green signal from target - ab58301 observed at 62 kDa
Lanes 3 and 4: Red signal from loading control - ab181602 observed at 37 kDa
Lanes 5 and 6: Merged (red and green) signal
ab58301 was shown to specifically react with PKR when PKR knockout samples were used. Wild-type and PKR knockout samples were subjected to SDS-PAGE. ab58301 and ab181602 (loading control to GAPDH) were diluted to 1 µg/mL and 1/2000 respectively and incubated overnight at 4°C. Blots were developed with Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) and Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) secondary antibodies at 1/10000 dilution for 1 h at room temperature before imaging.
ab58301 staining PKR in wild-type HAP1 cells (top panel) and EIF2AK2 knockout HAP1 cells (bottom panel). The cells were fixed with 100% methanol (5min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated with ab58301 at 1μg/ml concentration and ab6046 at 1μg/ml concentration overnight at +4°C, followed by a further incubation at room temperature for 1h with a goat secondary antibody to Mouse IgG (Alexa Fluor® 488) (ab150117) at 2 μg/ml (shown in green) and a goat secondary antibody to Rabbit IgG (Alexa Fluor® 594) (ab150084) at 2 μg/ml (showed in pseudocolour red) . Nuclear DNA was labelled in blue with DAPI.
ab58301 at 10 ug/ml staining PKR in human HeLa cells by Immunocytochemistry/ Immunofluorescence.