1/100 - 1/500. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Carbamoyl phosphate synthetase-aspartate carbamoyltransferase-dihydroorotase (CAD) is a multifunctional protein that initiates and regulates mammalian de novo pyrimidine biosynthesis. This trifunctional protein which is associated with the enzymatic activities of the first 3 enzymes in the 6-step pathway of pyrimidine biosynthesis is the rate-limiting step in the de novo pyrimidine synthetic pathway. Although most of the CAD protein in the cell is cytosolic, phosphorylation at threonine 456 localizes the protein to the nucleus. While MAPK and EGF phosphorylate CAD at threonine 456, MAPK and c-myc have been found to induce over-expression of CAD.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human colon carcinoma tissue labelling CAD with ab99311 at 1/200 (1µg/ml). Detection: DAB.
Western blot - Anti-CAD antibody (ab99311)
All lanes : Anti-CAD antibody (ab99311) at 0.04 µg/ml
Lane 1 : HeLa whole cell lysate at 50 µg Lane 2 : HeLa whole cell lysate at 15 µg Lane 3 : HeLa whole cell lysate at 5 µg
Predicted band size: 243 kDa
Exposure time: 10 seconds
Immunoprecipitation - Anti-CAD antibody (ab99311)
Detection of CAD by Western Blot of Immunprecipitate.
ab99311 at 0.1µg/ml staining CAD in HeLa whole cell lysate immunoprecipitated using ab99311 at 3µg/mg lysate (1 mg/IP; 20% of IP loaded/lane). Detection: Chemiluminescence with exposure time of 10 seconds.