ICC/IF image of ab107129 stained DU145 cells. The cells were 4% formaldehyde 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 ab107129 at 10µg/ml overnight at +4°C. The secondary antibody (green) was DyLight® 488 goat anti- chicken (ab96947) IgY (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.
IHC image of Mitofusin 1 staining in Human colon adenocarcinoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol B. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab107129, 5µg/ml, for 15 mins at room temperature. A Goat anti-Chicken biotinylated secondary antibody was used to detect the primary, and visualized using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
All lanes : Anti-Mitofusin 1 antibody (ab107129) at 1 µg/mlLane 1 : Kidney (Human) Tissue Lysate - adult normal tissue (ab30203)Lane 2 : Kidney (Mouse) Tissue Lysate Lane 3 : Kidney (Rat) Tissue LysateLane 4 : Heart (Human) Tissue Lysate - adult normal tissue (ab29431)Lane 5 : Heart (Mouse) Tissue Lysate Lane 6 : Heart (Rat) Tissue Lysate Lane 7 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate Lysates/proteins at 10 µg per lane.SecondaryGoat polyclonal Secondary Antibody to Chicken IgY - H&L (HRP) at 1/3000 dilutiondeveloped using the ECL techniquePerformed under reducing conditions.