Mitochondrial Pyruvate dehydrogenase kinase 1 was immunoprecipitated using 0.5mg Mouse Heart extract, 5µg of Rabbit polyclonal to Mitochondrial Pyruvate dehydrogenase kinase 1 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-). The antibody was incubated under agitation with Protein G beads for 10min, Mouse Heart extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation. Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab92959. Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697). Band: 47kDa: Mitochondrial Pyruvate dehydrogenase kinase 1.
ICC/IF image of ab92595 stained HepG2 cells. The cells were 100% methanol fixed (5 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 (ab92959, 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) HeLa cells at 1µg/ml.
IHC image of Mitochondrial Pyruvate dehydrogenase kinase 1 staining in human normal heart FFPE section, performed on a 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 ab92959, 10µg/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
References for Anti-Mitochondrial Pyruvate dehydrogenase kinase 1 antibody (ab92959)
This product has been referenced in:
Li YG et al. 1-deoxynojirimycin inhibits glucose absorption and accelerates glucose metabolism in streptozotocin-induced diabetic mice. Sci Rep3:1377 (2013).
Read more (PubMed: 23536174) »
Li YG et al. Cooperative anti-diabetic effects of deoxynojirimycin-polysaccharide by inhibiting glucose absorption and modulating glucose metabolism in streptozotocin-induced diabetic mice. PLoS One8:e65892 (2013).
Read more (PubMed: 23755289) »