The HTRF KinEASE assay series is a semi-universal biochemical platform to study kinase activity. The STK2 kit uses a specific substrate compatible with various Serine/thronine kinases - cf. validation data provided on this page.
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  • Low enzyme consumption Low enzyme consumption
The HTRF KinEASE assay series is a semi-universal biochemical platform to study kinase activity. The STK2 kit uses a specific substrate compatible with various Serine/thronine kinases - cf. validation data provided on this page.
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Overview

The HTRF KinEASE kit series provides a simple biochemical approach to study kinase activity, screen for inhibitors and characterize them. Three STK and one TK kits, offer a semi-universal method for assessing phosphorylation on Serine/Threonine and Tyrosine residues, respectively. The HTRF KinEASE STK S2 assay has been validated on 18 Serine/Threonine kinases. The assay validation section of this page lists them and provides technical tips for setup. The kit series also include a Discovery kit to help determine which of three STK kits is appropriate when working with a non-validated enzyme.

Benefits

  • COMPATIBLE WITH UP TO MM ATP KMS
  • SEMI-UNIVERSAL METHOD FOR STKS
  • LOW ENZYME CONSUMPTION

Assay principle

Each HTRF KinEASE kit is based on the same assay format, and uses a universal biotinylated substrate, a phospho-specific Eu-cryptate-labeled monoclonal antibody, and SA-XL665. All three STK peptidic substrates are made of the same sequence for antibody detection, next to a variable sequence recognized by enzymes. The targeted residue for phosphorylation lies in between as part of both sequences for enzymatic activity (unphosphorylated) and antibody detection (phosphorylated). It has been shown that 3 different variable sequences could enable the study of most Serine/Threonine kinases.

Assay protocol

As shown here, all HTRF KinEASE assays work the same way and involve two steps:

Step 1: Kinase reaction The kinase is incubated in the presence or absence of compounds and the appropriate substrate. The supplemented enzymatic buffer is added in the kinase buffer for tyr kinases only. ATP is added to start the reaction.

Step 2: Detection Eu-cryptate and XL665 conjugates are added. The detection buffer contains EDTA to stop the enzymatic reaction.

HTRF KinEASE Assay principle
Click in the table below to view the list of enzymes, the kit whose use is recommended, and to download the technical datasheet including experimental setting and validation data. Non listed STKs can be assessed with the Discovery kit to find out which HTRF KinEASE STK kit may be used.
ABL1 (TK)ABL2 (TK)AKT1 (STK S3)AKT2 (STK S3)AKT3 (STK S3)
ALK (TK)AMPK a2/ß1/g1 (STK S1)AMPK a2/ß2/g1 (STK S1)Arg (TK)ARK5 (STK S3)
ASK1 (STK S3)AURKA (STK S2)AURKB (STK S2)Aurora A (STK S2)Aurora B (STK S2)
Aurora C (STK S3)AXL (TK)BLK (TK)BMX (TK)BRK (TK)
BrSK1 (STK S3)BrSK2 (STK S1)BTK (TK)c-Kit (TK)CAMK1 (STK S1)
CaMK1 a (STK S1)CaMK2 a (STK S1)CaMK2 ß (STK S1)CaMK2 d (STK S1)CaMK2 g (STK S1)
CAMK2B (STK S1)CAMK2D (STK S1)CAMK2G (STK S1)CaMK4 (STK S1)CDC42 BPA (STK S3)
CDC42 BPB (STK S3)CGK2 (STK S2)CHEK1 (STK S1)CHEK2 (STK S1)CHK1 (STK S1)
CHK2 (STK S1)CHUK (STK S3)CLK3 (STK S3)COT (STK S3)CSF1R (TK)
CSK (TK)DAPK1 (STK S1)DAPK2 (STK S1)DAPK3 (STK S1)DCAMKL2 (STK S1)
DDR2 (TK)DMPK (STK S3)DRAK1 (STK S1)DYRK2 (STK S3)EEF-2K (STK S3)
EGFR (TK)EPHA1 (TK)EPHA2 (TK)EPHA3 (TK)EPHA4 (TK)
EPHA5 (TK)EPHA7 (TK)EPHA8 (TK)EPHB1 (TK)EPHB2 (TK)
EPHB3 (TK)EPHB4 (TK)ERBB4 (TK)FAK (TK)FER (TK)
FES (TK)FGFR1 (TK)FGFR2 (TK)FGFR3 (TK)FGFR4 (TK)
FGR (TK)FLT1 (TK)FLT3 (TK)FLT4 (TK)FMS (TK)
Fps (TK)FRK (TK)FYN (TK)GPRK4 (STK S1)GRK4 (STK S1)
GRK5 (STK S3)GRK6 (STK S1)GSG2 (STK S1)GSK3 a (STK S3)Haspin (STK S1)
HCK (TK)HER4 (TK)HIPK2 (STK S3)HIPK3 (STK S3)IGF1R (TK)
IKBKB (STK S3)IKK a (STK S3)IKK ß (STK S3)INSR (TK)INSRR (TK)
Insulin R (TK)IRAK4 (STK S1)IRR (TK)ITK (TK)JAK1 (TK)
JAK2 (TK)JAK3 (TK)KDR (TK)KIT (TK)LATS1 (STK S1)
LCK (TK)LKB1 (STK S1)LOK (STK S3)Lyn (TK)LYNa (TK)
MAP3K8 (STK S3)MAPKAP-K1 a (STK S1)MAPKAP-K1 ß (STK S1)MAPKAP-K2 (STK S1)MAPKAP-K3 (STK S1)
MAPKAP-K5 (STK S1)MARK1 (STK S1)MARK2 (STK S2)MELK (STK S1)MER (TK)
MERTK (TK)MET (TK)MINK1 (STK S3)MKNK1 (STK S1)MKNK2 (STK S1)
MLCK (STK S1)MNK1 (STK S1)MNK2 (STK S1)MRCK a (STK S3)MRCK ß (STK S3)
MSK1 (STK S3)MSK2 (STK S3)MSSK1 (STK S3)MST1 (STK S3)MST1R (TK)
MST2 (STK S3)MSTIR (TK)MUSK (TK)MYLK (STK S1)NEK11 (STK S2)
NEK2 (STK S3)NEK3 (STK S3)NEK6 (STK S3)NEK7 (STK S3)NLK (STK S3)
NTRK1 (TK)NTRK2 (TK)NTRK3 (TK)p70S6K (STK S3)PAK2 (STK S2)
PAK3 (STK S2)PAK4 (STK S2)PAK5 (STK S2)PAK6 (STK S2)PAK7 (STK S2)
PAR-1B a (STK S2)PASK (STK S1)PDGFR a (TK)PDGFR ß (TK)PDGFRA (TK)
PDGFRB (TK)PHK g1 (STK S1)PHK g2 (STK S1)PIM1 (STK S3)PIM2 (STK S3)
PKAC a (STK S2)PKAC g (STK S3)PKB a (STK S3)PKB ß (STK S3)PKB g (STK S3)
PKC a (STK S1)PKC ß1 (STK S1)PKC ß2 (STK S1)PKC d (STK S1)PKC epsilon (STK S1)
PKC µ (STK S1)PKC theta (STK S1)PKC eta (STK S1)PKC gamma (STK S1)PKC iota (STK S1)
PKC zeta (STK S1)PKD2 (STK S1)PKG1 a (STK S2)PKG1 ß (STK S2)PKG2 (STK S1)
PKN2 (STK S1)PLK1 (STK S1)PLK3 (STK S3)PLK4 (STK S2)PRAK (STK S1)
PRK2 (STK S1)PRKA A2/B1/G1 (STK S1)PRKACA (STK S2)PRKACG (STK S3)PRKCA (STK S1)
PRKCB1 (STK S1)PRKCB2 (STK S1)PRKCD (STK S1)PRKCE (STK S1)PRKCG (STK S1)
PRKCH (STK S1)PRKCI (STK S1)PRKCQ (STK S1)PRKCZ (STK S1)PRKD2 (STK S1)
PRKG1 (STK S2)PRKG2 (STK S1)PRKX (STK S2)PTK2 (TK)PTK2B (TK)
PTK5 (TK)PTK6 (TK)PYK2 (TK)RET (TK)ROCK1 (STK S2)
ROCK2 (STK S2)ROK a (STK S2)RON (TK)ROS (TK)ROS1 (TK)
RPS6KA1 (STK S1)RPS6KA2 (STK S1)RPS6KA3 (STK S1)RPS6KA4 (STK S3)RPS6KA5 (STK S3)
RPS6KA6 (STK S2)RPS6KB1 (STK S3)Rse (TK)RSK1 (STK S1)RSK2 (STK S1)
RSK3 (STK S1)RSK4 (STK S2)SAD1 (STK S3)SGK1 (STK S3)SGK2 (STK S3)
SGK3 (STK S3)SGKL (STK S3)SIK (STK S1)smMLCK (STK S1)Snk (STK S3)
SRC (TK)STK22B (STK S1)STK22D (STK S1)STK23 (STK S3)STK3 (STK S3)
STK4 (STK S3)STK6 (STK S2)SYK (TK)TBK1 (STK S1)TEK (TK)
TIE2 (TK)TRKA (TK)TRKB (TK)TRKC (TK)TSSK1 (STK S1)
TSSK2 (STK S1)TYK2 (TK)TYR03 (TK)TYRO3 (TK)VEGFR1 (TK)
VEGFR2 (TK)VEGFR3 (TK)WNK2 (STK S3)WNK3 (STK S3)YES1 (TK)
ZAP70 (TK)ZIPK (STK S1)

Validated kinases

Over 190 Serine/Threonine and Tyrosine kinases have already been validated with the HTRF KinEASE kits. Out of those, 18 were found to be compatible with the S2 kit. The case of ROCK2 Serine/Threonine kinase is illustrated below with the enzyme titration and time course (on the left), and the inhibition by staurosporine (on the right).
ROCK2 kinase titration and time course
ROCK2 kinase inhibition by staurosporine
Aurora-A, AURKA, STK6 validated with HTRF KinEASE

Enzyme validation - Technical Notes

Aurora-B, AURKB validated with HTRF KinEASE

Enzyme validation - Technical Notes

CGK2, PRKG2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PAK2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PAK3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PAK4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PAK5, PAK7 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PAK6 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PKAC-alpha, PRKACA validated with HTRF KinEASE

Enzyme validation - Technical Notes

PKG1-alpha, PRKG1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PLK4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

PRKX validated with HTRF KinEASE

Enzyme validation - Technical Notes

ROCK1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

ROCK2, ROK-alpha validated with HTRF KinEASE

Enzyme validation - Technical Notes

RSK4, RPS6KA6 validated with HTRF KinEASE

Enzyme validation - Technical Notes

HTRF KinEASE

Universal platform to address Ser/Thr & Tyr kinases - Application Notes

Open R&D: Sanofi Access Platform

In collaboration with Sanofi - Scientific Presentations

Progress towards a generic intracellular kinase assay

In collaboration with Genomics institute of the Novartis Research Foundation - Scientific Presentations

KinEASE, a universal expanded platform to adress Ser/Thr & Tyr kinases

Measuring kinase activity under any conditions - Application Notes

Exploration is easier with a trusted partner

HTRF KinEASE Tyr and Ser/Thr kits - Flyers

Cisbio KinEASE

Validated kinases and assay conditions - Flyers

Product Insert KinEASE STK S2 Kit / 62ST2PEB-62ST2PEC-62ST2PEJ

62ST2PEB-62ST2PEC-62ST2PEJ - Product Insert

Cisbio Product Catalog 2019

All your HTRF assays in one document! - Catalog

A guide to Homogeneous Time Resolved Fluorescence

General principles of HTRF - Guides

The essential white paper to expand your knowledge on kinases

Learn about cutting edge kinase research in this free white paper - Guides

How HTRF compares to Western Blot and ELISA

Get the brochure about technology comparison. - Brochures

Multiple therapeutic areas adressed with HTRF KinEase assays

Discover how peer researchers challenge kinase investigations with KinEase assays - Application Notes

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