Abcam is committed to meeting high standards of ethical manufacturing and as such, we will be discontinuing this product, which has been generated by the ascites method, within the next year. We are sorry for any inconvenience this may cause. If you would like help finding an alternative product, please do not hesitate to contact our scientific support team.
Our Abpromise guarantee covers the use of ab57644 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: 16 kDa.|
|Flow Cyt||Use 1µg for 106 cells.
ab170191 - Mouse monoclonal IgG2a, is suitable for use as an isotype control with this antibody.
|ICC/IF||Use a concentration of 10 µg/ml.|
ab57644 staining RPS23 (right) compared to control IgG (left) in U2OS cells by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with 0.1% Triton X-100 and blocked with 2% BSA for 1 hour at 21°C. Samples were incubated with the primary antibody (2.5µg/ml in PBS + 1% BSA) for 16 hours at 4°C. An Alexa Fluor® 488-conjugated donkey anti-mouse IgG polyclonal (1/1000) was used as the secondary antibody. Counterstained with DAPI.
Overlay histogram showing HeLa cells stained with ab57644 (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 (ab57644, 1µg/1x106 cells) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was mouse IgG2a [ICIGG2A] (ab91361, 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.
ICC/IF image of ab57644 stained HeLa cells. The cells were 4% formaldehyde (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 (ab57644, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab96879 Dylight 488 goat anti-mouse IgG (H+L) used at a 1/250 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.