Synthetic phosphopeptide corresponding to residues surrounding S729 of human PKC.
Species Reactivity
Human, Mouse, Rat
Validated Applications
WB, IHC-P
Target Molecular Weight
Predicted band size: 84 kDa
Positive Control
HeLa cell lysate, HeLa treated with 200nM TPA for 30 minutes cell lysate, C2C12 cell lysate, C2C12 treated with 200nM TPA for 30 minutes cell lysate, C6 cell lysate, C6 treated with 200nM TPA for 10 minutes cell lysate, human breast cancer tissue.
Conjugation
unconjugated
Clone Number
JE77-54
Product Features
Form
Liquid
Concentration
1 mg/mL.(The concentration of this product may be batch-dependent)
Lot Concentration Lookup
Storage Instructions
Shipped at 4℃. Store at +4℃ short term (1-2 weeks). Store at -20℃ long term.
In cell biology, Protein kinase C, commonly abbreviated to PKC (EC 2.7.11.13), is a family of protein kinase enzymes that are involved in controlling the function of other proteins through the phosphorylation of hydroxyl groups of serine and threonine amino acid residues on these proteins, or a member of this family. PKC enzymes in turn are activated by signals such as increases in the concentration of diacylglycerol (DAG) or calcium ions (Ca2+). Hence PKC enzymes play important roles in several signal transduction cascades. In biochemistry, the PKC family consists of fifteen isozymes in humans. They are divided into three subfamilies, based on their second messenger requirements: conventional (or classical), novel, and atypical. Conventional (c)PKCs contain the isoforms α, βI, βII, and γ. These require Ca2+, DAG, and a phospholipid such as phosphatidylserine for activation. Novel (n)PKCs include the δ, ε, η, and θ isoforms, and require DAG, but do not require Ca2+ for activation. Thus, conventional and novel PKCs are activated through the same signal transduction pathway as phospholipase C. On the other hand, atypical (a)PKCs (including protein kinase Mζ and ι / λ isoforms) require neither Ca2+ nor diacylglycerol for activation.
Background References
1. Lin G, Brownsey R W, MacLeod K M. Complex regulation of PKCβ2 and PDK-1/AKT by ROCK2 in diabetic heart[J]. PLoS One, 2014, 9(1): e86520.
2. Ghosh P M, Bedolla R, Thomas C A, et al. Role of protein kinase C in arginine vasopressin‐stimulated ERK and p70S6 kinase phosphorylation[J]. Journal of cellular biochemistry, 2004, 91(6): 1109-1129.
Western blot analysis of Phospho-PKC epsilon (S729) on different lysates with Rabbit anti-Phospho-PKC epsilon (S729) antibody (HA722846) at 1/1,000 dilution.
Lane 1: HeLa cell lysate Lane 2: HeLa treated with 200nM TPA for 30 minutes cell lysate Lane 3: C2C12 cell lysate Lane 4: C2C12 treated with 200nM TPA for 30 minutes cell lysate Lane 5: C6 cell lysate Lane 6: C6 treated with 200nM TPA for 10 minutes cell lysate Lane 7: HeLa treated with 200nM TPA for 30 minutes cell lysate, then the membrane treated with Lambda PP for 1 hour Lane 8: C2C12 treated with 200nM TPA for 30 minutes cell lysate, then the membrane treated with Lambda PP for 1 hour Lane 9: C6 treated with 200nM TPA for 10 minutes cell lysate, then the membrane treated with Lambda PP for 1 hour
Lysates/proteins at 20 µg/Lane.
Predicted band size: 84 kDa Observed band size: 84 kDa
Exposure time: 59 seconds; ECL: K1802;
4-20% SDS-PAGE gel.
Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA722846) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
Immunohistochemical analysis of paraffin-embedded human breast cancer tissue with Rabbit anti-Phospho-PKC epsilon (S729) antibody (HA722846) at 1/200 dilution.
The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA722846) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
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