The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
1/2000 - 1/10000. Detects a band of approximately 47 kDa.
Use at an assay dependent concentration.
1/250 - 1/500.
ab172730 - Rabbit monoclonal IgG, is suitable for use as an isotype control with this antibody.
Adapter protein which associates with tyrosine-phosphorylated growth factor receptors or their cellular substrates. Maintains low levels of EIF2S1 phosphorylation by promoting its dephosphorylation by PP1. Plays a role in the DNA damage response, not in the detection of the damage by ATM/ATR, but for efficient activation of downstream effectors, such as that of CHEK2.
Contains 1 SH2 domain. Contains 3 SH3 domains.
Phosphorylated on Ser and Tyr residues.
Cytoplasm. Endoplasmic reticulum. Nucleus. Mostly cytoplasmic, but shuttles between the cytoplasm and the nucleus. Import into the nucleus requires the interaction with SOCS7. Predominantly nuclear following genotoxic stresses, such as UV irradiation, hydroxyurea or mitomycin C treatments.
Overlay histogram showing Jurkat cells stained with ab32120 (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 (ab32120, 1/1000 dilution) for 30 min at 22°C. The secondary antibody used was Alexa Fluor® 488 goat anti-rabbit IgG (H&L) (ab150077) at 1/2000 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit IgG (monoclonal) (0.1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >5,000 events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. This antibody gave a positive signal in Jurkat cells fixed with 80% methanol (5 min)/permeabilized with 0.1% PBS-Tween for 20 min used under the same conditions.
Western blot - Nck antibody [Y531] (ab32120)
Anti-Nck antibody [Y531] (ab32120) at 1/10000 dilution + Hela cell lysate