The antibody has been negatively preadsorbed using a non-phosphopeptide corresponding to the site of phosphorylation to remove antibody that is reactive with non-phosphorylated PKC eta. The final product is generated by affinity chromatography using a PKC eta-derived peptide that is phosphorylated at threonine 655.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 0.35 - 1 µg/ml. Detects a band of approximately 80 kDa.
This is calcium-independent, phospholipid-dependent, serine- and threonine-specific enzyme. PKC is activated by diacylglycerol which in turn phosphorylates a range of cellular proteins. PKC also serves as the receptor for phorbol esters, a class of tumor promoters.
Most abundant in lung, less in heart and skin.
Defects in PRKCH may be a cause of susceptibility to ischemic stroke (ISCHSTR) [MIM:601367]; also known as cerebrovascular accident or cerebral infarction. A stroke is an acute neurologic event leading to death of neural tissue of the brain and resulting in loss of motor, sensory and/or cognitive function. Ischemic strokes, resulting from vascular occlusion, is considered to be a highly complex disease consisting of a group of heterogeneous disorders with multiple genetic and environmental risk factors.
Belongs to the protein kinase superfamily. AGC Ser/Thr protein kinase family. PKC subfamily. Contains 1 AGC-kinase C-terminal domain. Contains 1 C2 domain. Contains 2 phorbol-ester/DAG-type zinc fingers. Contains 1 protein kinase domain.
The C1 domain, containing the phorbol ester/DAG-type region 1 (C1A) and 2 (C1B), is the diacylglycerol sensor and the C2 domain is a non-calcium binding domain.
Western blot - Anti-PKC eta (phospho T655) antibody (ab5798)
Peptide Competition and Phosphatase Treatment Lysates prepared from Jurkat cells stimulated with PMA were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were either left untreated (1-9) or treated with lambda phosphatase (10), blocked with a 5% BSA-TBST buffer overnight at 4°C, and incubated with 0.50 µg/mL ab5798 antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 10), the non phosphopeptide corresponding to the immunogen (2), a generic phosphothreonine containing peptide (3), the phosphopeptide immunogen (4), or, the phosphopeptide corresponding to the immunogen from other PKC isoforms (5-9). After washing, membranes were incubated with goat (ab’)2 anti-rabbit IgG HRP-conjugate and bands were detected using the Pierce SuperSignalTM method. The data show that the peptide corresponding to PKC eta [pT655] blocks the antibody signal. The antibody signal was not blocked by the peptides corresponding to PKC isoforms alpha [pT638], beta 1 [pT642], beta 1 [pT641], gamma [pT655] and zeta [pT560], thereby demonstrating the specificity of the antibody. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific.