Overview

  • Product name

    PKC Kinase Activity Assay Kit
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Sample < 10%
    Inter-assay
    Sample n Mean SD CV%
    Sample < 10%
  • Sample type

    Adherent cells, Suspension cells, Tissue Extracts, Purified protein
  • Assay type

    Enzyme activity
  • Assay time

    4h 30m
  • Product overview

    PKC Kinase Activity Assay Kit (ab139437) is a non-radioactive assay providing a safe, rapid and reliable method for the screening of inhibitors or activators of PKC and for quantitating the activity of PKC in purified or partially purified enzyme preparations. This kit is based on a solid phase enzyme-linked immuno-absorbent assay (ELISA) that utilizes a specific synthetic peptide as a substrate for PKC and a polyclonal antibody that recognizes the phosphorylated form of the substrate. The assay is designed for the analysis of PKC activity in the solution phase. For the measurement of PKC in partially purified, purified, or crude enzyme preparations from any species.

    The kit offers the following advantages:

    1. Safe - non-radioactive measurement of kinase activity.

    2. Flexible - kinetic and end-point options available.

    3. Fast - results in < 4.5 hours.

    4. Efficient - only 30 μl diluted sample needed per well.

  • Platform

    Microplate reader

Properties

  • Storage instructions

    Please refer to protocols.
  • Components 1 x 96 tests
    20X Wash Buffer 1 x 30ml
    Active PKC 1 x 28µl
    Antibody Dilution Buffer 1 x 10ml
    Anti-Rabbit IgG: HRP Conjugate 1 x 20µl
    ATP 1 x 2mg
    Kinase Assay Dilution Buffer 1 x 10ml
    PKC Phosphospecific Substrate Antibody 1 x 5ml
    PKC Substrate Microtiter Plate 1 unit
    Stop Solution 2 1 x 10ml
    TMB Substrate 1 x 10ml
  • Research areas

  • Relevance

    Protein Kinase C (PKC) isoforms are serine/threonine kinases involved in signal transduction pathways that govern a wide range of physiological processes including differentiation, proliferation, gene expression, brain function, membrane transport and the organization of cytoskeletal and extracellular matrix proteins. Increasing evidence from studies using in vitro and in vivo systems points to PKC as a key regulator of critical cell cycle transitions, including cell cycle entry and exit and the G1 and G2 checkpoints. PKC-mediated control of these transitions can be negative or positive, depending on the timing of PKC activation during the cell cycle and on the specific PKC isozymes involved. There have been at least 12 different PKC isoforms identified in humans to date, including alpha, beta I, beta II, gamma, delta, epsilon, zeta, eta, theta, iota, lambda, and mu.

Images

  • 1.5 x e7 THP-1 cells were incubated with 100 nM GF109203X (GF; ab144264) or Ro31-8220 mesylate (Ro; ab120374) for 30 minutes prior to activation with 10 μg x mL-1 PMA (Sigma) for 4 hours. Control cells were left without inhibitors or PMA. Cells were lysed in 1 mL of lysis buffer, and 30 μL were tested for PKC activity (duplicates; +/- SD).

  • Titration of ab139437 (duplicates; +/- SD).

  • Signal from active PKC with background signal subtracted (duplicates; +/- SD).

Protocols

References

This product has been referenced in:

  • Koyyada R  et al. Naltrexone Reverses Ethanol Preference and Protein Kinase C Activation in Drosophila melanogaster. Front Physiol 9:175 (2018). Functional Studies . Read more (PubMed: 29593550) »
  • Asensio-Lopez MDC  et al. Pharmacological inhibition of the mitochondrial NADPH oxidase 4/PKCa/Gal-3 pathway reduces left ventricular fibrosis following myocardial infarction. Transl Res N/A:N/A (2018). Read more (PubMed: 29753686) »
See all 12 Publications for this product

Customer reviews and Q&As

1-3 of 3 Abreviews or Q&A

Question
Answer

It is possible that the lysis buffer may work, but we can't guarantee it. I would suggest comparing your reagents to our components in our lysis buffer and try to add any vital or missing components especially the inhibitors that need to be added fresh before lysis.

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Question
Answer

The concentration of the active PKC is 20ng/ul for every lot.

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Answer


1) Which PKC isoforms are being measured with this kit?

The kit is not specific for any one isoform; the kit is merely dependent on sufficient levels of active PKC. The active PKC, regardless of the isoforms present, will drive phosphorylation to the plate during incubation with the ATP.


2) Was the kit also tested on a cell line? If so, which one?

The kit was not tested on a cell line and was only tested using a purified PKC, active enzyme. Many researchers will use this kit for testing samples such as cell lysates; they need to be sure to optimize dilution and/or concentration of that sample to add to the assay as well as optimize incubation time for the enzymatic behavior of the active PKC present in their samples will need to be characterized to insure that the best parameters are used for their assay.


3) The kit was tested with which inhibitors?

The kit was not tested with any known PKC inhibitors or other inhibitors of any kind. However, we know that other researchers who have used this kit for inhibition studies.

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Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
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