All lanes : Anti-HEF1 antibody (ab37161) at 1/250 dilutionLane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell LysateLane 2 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate (ab7899)Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate with Human HEF1 peptide (ab39117) at 1 µg/mlLane 4 : Jurkat (Human T cell lymphoblast-like cell line) Whole Cell Lysate (ab7899) with Human HEF1 peptide (ab39117) at 1 µg/mlLysates/proteins at 10 µg per lane.SecondaryIRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilutionPerformed under reducing conditions.
ab37161 staining HEF1 in Human glioblastoma cells by Immunocytochemistry/ Immunofluorescence. Cells were PFA-fixed and permeabilized in 0.1% Triton X-100 in PBS prior to blocking in 0.5% BSA in TBS-Tween for 20 minutes at room temperature. The primary antibody was diluted 1/50 in 0.5% BSA/PBS and incubated with the sample for 16 hours at 4°C. The secondary antibody was Cy3®-conjugated Goat anti-Rabbit IgG, diluted 1/400. Nuclei were counterstained with Hoechst.See Abreview
ICC/IF image of ab37161 stained HeLa cells. The cells were 4% PFA fixed (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 (ab37161, 5µg/ml) overnight at +4°C. The secondary antibody (green) was ab96899 Dylight® 488 goat anti-rabbit 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.