The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use at an assay dependent concentration. PubMed: 22532868
1/1000. Detects a band of approximately 82 kDa.
Use 5 - 10 µl.
1/100. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
This is a calcium-activated, phospholipid-dependent, serine- and threonine-specific enzyme. May play a role in cell motility by phosphorylating CSPG4. 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.
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.
Western blot - Anti-PKC alpha (phospho S657 + Y658) antibody (ab23513)
Western blot analysis of immunoprecipitates from neonatal rat brain lysate using anti-PKCα antibody. Control and alkaline phosphatase treated precipitates were probed with anti-PKCα (Central region) or anti-phospho-PKCα (Ser-657/Tyr-658).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PKC alpha (phospho S657 + Y658) antibody (ab23513)This image is courtesy of an anonymous Abreview
ab23513 staining PKC alpha (phospho S657 + Y658) in human pancreatic ductal adenocarcinoma tissue sections by Immunohistochemistry (IHC-P - formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde, permeabilized with PBS + Triton 0.025% and blocked with 5% serum for 1 hour at room temperature; antigen retrieval was by heat mediation with a citrate buffer (pH 6). Samples were incubated with primary antibody (1/100 in PBS) for 8 hours at 4°C. A Biotin-conjugated Goat anti-rabbit polyclonal (1/1000) was used as the secondary antibody.
Immunocytochemical labeling of PKC phosphorylation in aldehyde-fixed and NP-40-permeabilized NGF-differentiated PC12 cells. The cells were labeled with rabbit polyclonal anti-PKCα (Ser-657/Tyr-658) (PP1091) antibody in the absence (Left) or presence (Right) of blocking peptide (PX1095). The antibody was detected using appropriate secondary antibody conjugated to DyLight® 594
Immunocytochemistry/ Immunofluorescence - Anti-PKC alpha (phospho S657 + Y658) antibody (ab23513)Image from Taniuchi K et al., PLoS One. 2012;7(4):e35674. Epub 2012 Apr 19. Fig 8.; doi:10.1371/journal.pone.0035674; April 19, 2012, PLoS ONE 7(4): e35674.
Immunofluorescence analysis of S2-013 cells, staining PKC alpha (phospho S657 + Y658) with ab23513.
Cells were fixed with paraformaldehyde, permeabilized with 0.1% Triton X-100, and blocked with blocking solution (3% BSA/PBS). Cells were incubated with primary antibody for 1 hour. An AlexaFluor®488-conjugated anti-rabbit IgG was used as the secondary antibody.
Wang S et al. Regulation of endothelial cell proliferation and vascular assembly through distinct mTORC2 signaling pathways. Mol Cell Biol35:1299-313 (2015).
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