The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Detects a band of approximately 9 kDa (predicted molecular weight: 11 kDa).
Use a concentration of 1 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.
At least 9 distinct polymorphic forms of apolipoproteins are known. The apolipoproteins act as stabilizers of the intact lipoprotein particles. Quantitative measurements of HDL, LDL and VLDL particles in human serum are often used to estimate an individuals' relative risk of coronary heart disease. In addition, quantitative immunological measurements of certain apolipoproteins (especially A-1 and B) have been suggested to be more accurate estimators of coronary heart disease than measurements of lipoprotein particles (especially HDL and LDL).
Apolipoprotein C-II (apoCII) is in found in chylomicrons (large lipoprotein particles absorbed from the gastrointestinal tract) and VLDL (large lipoproteins that are broken down to eventually form LDL). ApoCII activates the enzyme lipoprotein lipase, which hydrolyzes triglycerides and thus provides free fatty acids for cells.
The Apolipoprotein CII protein has a predicted molecular weight of 11-kDa. The first 22 amino-acids of the Apolipoprotein CII sequence act as a signal sequence, and when cleaved the protein has an expected molecular weight of 9-kDa.
ICC/IF image of ab76452 stained HeLa 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 (ab76452, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit 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) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) HepG2 and MCF7 cells at 1µg/ml, and in 100% methanol fixed (5 min) HeLa cells at 1µg/ml
IHC image of Apolipoprotein CII staining in human normal liver formalin fixed paraffin embedded tissue section, performed on a Leica BondTM 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 ab76452, 1µg/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.
Okubo M et al. Apolipoprotein C-II Tuzla: a novel large deletion in APOC2 caused by Alu-Alu homologous recombination in an infant with apolipoprotein C-II deficiency. Clin Chim Acta438:148-53 (2015).
Read more (PubMed: 25172036) »