The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Use a concentration of 1 µg/ml. Detects a band of approximately 14 kDa (predicted molecular weight: 14 kDa).
Lipid transport protein in adipocytes. Binds both long chain fatty acids and retinoic acid. Delivers long-chain fatty acids and retinoic acid to their cognate receptors in the nucleus.
Belongs to the calycin superfamily. Fatty-acid binding protein (FABP) family.
Forms a beta-barrel structure that accommodates hydrophobic ligands in its interior.
Cytoplasm. Nucleus. Depending on the nature of the ligand, a conformation change exposes a nuclear localization motif and the protein is transported into the nucleus. Subject to constitutive nuclear export.
IHC image of FABP4 staining in human lung formalin fixed paraffin embedded tissue section, performed on a Leica Bond system using the standard protocol F. 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 ab85875, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer 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.
Western blot - FABP4 antibody (ab85875)
All lanes : Anti-FABP4 (phospho Y20) antibody (ab85875) at 1 µg/ml
Lane 1 : Human adipose normal tissue lysate - total protein (ab28980) Lane 2 : Adult Rat Brown Adipose Tissue Tissue Lysate
Lysates/proteins at 10 µg per lane.
Secondary All lanes : Goat Anti-Rabbit IgG H&L (HRP) preadsorbed (ab97080) at 1/5000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 14 kDa Observed band size: 14 kDa
Exposure time: 16 minutes
Western blot - Anti-FABP4 antibody (ab85875)
Anti-FABP4 (phospho Y20) antibody (ab85875) at 1 µg/ml + Adult Mouse Brown Adipose Tissue Tissue Lysate at 10 µg