Anti-ATPB antibody [3D5] - Mitochondrial Marker (ab14730)

  Immunocytochemistry/ Immunofluorescence - ATPB antibody [3D5] (ab14730)

ICC/IF image of ab14730 stained human HepG2 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 (ab14730, 5 µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 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). This antibody also gave a positive IF result in HeLa, Hek293 and MCF7 cells.

  Western blot - ATPB antibody [3D5] (ab14730)

All lanes : Anti-ATPB antibody [3D5] - Mitochondrial Marker (ab14730) at 0.8 µg/ml

Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate
Lane 2 : HepG2 (Human hepatocellular liver carcinoma cell line) Whole Cell Lysate
Lane 3 : Liver (Human) Tissue Lysate - adult normal tissue (ab29889)

Lysates/proteins at 10 µg per lane.

Secondary
Goat polyclonal to Mouse IgG - H&L - Pre-Adsorbed (HRP) at 1/3000 dilution

Predicted band size : 52 kDa
Observed band size : 52 kDa

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - ATPB antibody [3D5] (ab14730)

ab14730 (2µg/ml) staining ATPB in human duodenum using an automated system (DAKO Autostainer Plus). Using this protocol there is cytoplasmic and mitochondrial staining of epithelium.
Sections were rehydrated and antigen retrieved with the Dako 3 in 1 AR buffer EDTA pH 9.0 in a DAKO PT link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 mins. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 min and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with Diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that, for manual staining, optimization of primary antibody concentration and incubation time is recommended. Signal amplification may be required.

  Western blot - ATPB antibody [3D5] (ab14730)

All lanes : Anti-ATPB antibody [3D5] - Mitochondrial Marker (ab14730)

Lane 1 : isolated mitochondria from human heart at 5 µg
Lane 2 : isolated mitochondria from bovine heart at 1 µg
Lane 3 : isolated mitochondria from rat heart at 10 µg
Lane 4 : isolated mitochondria from mouse heart at 10 µg


Predicted band size : 52 kDa
Observed band size : 52 kDa

  Immunocytochemistry/ Immunofluorescence - ATPB antibody [3D5] (ab14730)

ab14730 staining ATPB in cultured human fibroblasts. Cells were fixed, permeabilized and then labelled with ab14730 followed by an AlexaFluor® 488-conjugated Goat anti-Mouse IgG1-specific secondary antibody (2 µg/ml)

  Flow Cytometry - ATPB antibody [3D5] (ab14730)

ab14730 (blue) at 1µg/ml staining ATPB in HL-60 cells and analyzed by Flow cytometry. Red histogram represents equal quantity of isotype control.