All lanes : Anti-Vasopressin V1b receptor antibody (ab104365) at 1 µg/mlLane 1 : E10 Mouse Embryo Brain Tissue Lysate Lane 2 : Forebrain (Mouse) Tissue LysateLane 3 : Brain (Human) Membrane Lysate - adult normal tissue (ab29456)Lane 4 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell LysateLane 5 : E10 Mouse Embryo Brain Tissue Lysate with Immunising peptide at 1 µg/mlLane 6 : Forebrain (Mouse) Tissue Lysate with Immunising peptide at 1 µg/mlLane 7 : Brain (Human) Membrane Lysate - adult normal tissue (ab29456) with Immunising peptide at 1 µg/mlLane 8 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate with Immunising peptide at 1 µg/mlLysates/proteins at 10 µg per lane.SecondaryGoat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.
ICC/IF image of ab104365 stained SKNSH 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 (ab104365, 1µg/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. This antibody also gave a positive result in 4% formaldehyde fixed (10 min) SKNSH cells at 1µg/ml.