ab75659 was affinity-purified from rabbit antiserum by affinity-chromatography using epitope-specific phosphopeptide. The antibody against non-phosphopeptide was removed by chromatography using non-phosphopeptide corresponding to the phosphorylation site.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 5 µg/ml.
1/100 - 1/200.
1/500 - 1/1000. Detects a band of approximately 148 kDa (predicted molecular weight: 148 kDa).
The production of the second messenger molecules diacylglycerol (DAG) and inositol 1,4,5-trisphosphate (IP3) is mediated by activated phosphatidylinositol-specific phospholipase C enzymes. It is a crucial enzyme in transmembrane signaling.
PLCG 2 was immunoprecipitated using 0.5mg A431 whole cell extract, 5µg of Rabbit polyclonal to PLCG 2 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, A431 whole cell 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 70°C; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab75659.