Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human ovarian carcinoma tissue labelling RAI1 with ab86599 at 1/1000 (1µg/ml). Detection: DAB.
ICC/IF image of ab86599 stained HeLa cells. The cells were 100% methanol fixed (5 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 ab86599 at 5µg/ml overnight at +4°C. The secondary antibody (pseudo-colored green) was Alexa Fluor® 488 goat anti- rabbit (ab150081) IgG (H+L) preadsorbed, used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (pseudo-colored red) at a 1/200 dilution for 1h at room temperature. DAPI was used to stain the cell nuclei (pseudo-colored blue) at a concentration of 1.43µM for 1hour at room temperature.
All lanes : Anti-RAI1 antibody (ab86599) at 0.1 µg/mlLane 1 : Whole cell lysate from HeLa cells at 50 µgLane 2 : Whole cell lysate from HeLa cells at 15 µgLane 3 : Whole cell lysate from HeLa cells at 5 µgdeveloped using the ECL technique
Western Blot detection of RAI1 in Immunoprecipitates of Whole cell lysate of HeLa cells (1 mg for IP, 20% of IP loaded) using ab86599 at 10 µg/mg lysate for IP and at 1 µg/ml for subsequent Western blot detection. Lane 2 represents control IgG IP. Detection: Chemiluminescence with an exposure time of 30 seconds.