The HTRF KinEASE assay series is a semi-universal biochemical platform to study kinase activity. The TK kit uses a specific substrate compatible with tyrosine kinases - cf. validation data provided on this page.
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The HTRF KinEASE assay series is a semi-universal biochemical platform to study kinase activity. The TK kit uses a specific substrate compatible with tyrosine 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 TK assay has been validated on 71 Tyrosine kinases, using a single generic substrate. The assay validation section of this page lists them and provides technical tips for setup.

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. The TK generic substrate is made of a sequence of repetitive motifs recognized by most tyrosine kinases, and by the specific conjugated antibody once phosphorylated.

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.

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.
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, 71 were tyrosine kinases that could be assessed with the TK kit. The case of PDGFRB Tyrosine kinase is illustrated below with the enzyme titration and time course (on the left), and the inhibition by staurosporine (on the right).
ABL1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

ABL2, Arg validated with HTRF KinEASE

Enzyme validation - Technical Notes

ALK validated with HTRF KinEASE

Enzyme validation - Technical Notes

AXL validated with HTRF KinEASE

Enzyme validation - Technical Notes

BLK validated with HTRF KinEASE

Enzyme validation - Technical Notes

BMX validated with HTRF KinEASE

Enzyme validation - Technical Notes

BRK, PTK6 validated with HTRF KinEASE

Enzyme validation - Technical Notes

BTK validated with HTRF KinEASE

Enzyme validation - Technical Notes

CSK validated with HTRF KinEASE

Enzyme validation - Technical Notes

DDR2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EGFR validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA5 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA7 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHA8 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHB1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHB2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHB3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

EPHB4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FAK, PTK2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FER validated with HTRF KinEASE

Enzyme validation - Technical Notes

FES, Fps validated with HTRF KinEASE

Enzyme validation - Technical Notes

FGFR1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FGFR2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FGFR3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FGFR4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FGR validated with HTRF KinEASE

Enzyme validation - Technical Notes

FLT1, VEGFR1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FLT3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FLT4, VEGFR3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FMS, CSF1R validated with HTRF KinEASE

Enzyme validation - Technical Notes

FRK, PTK5 validated with HTRF KinEASE

Enzyme validation - Technical Notes

FYN validated with HTRF KinEASE

Enzyme validation - Technical Notes

HCK validated with HTRF KinEASE

Enzyme validation - Technical Notes

HER4, ERBB4 validated with HTRF KinEASE

Enzyme validation - Technical Notes

IGF1R validated with HTRF KinEASE

Enzyme validation - Technical Notes

INSR, Insulin R validated with HTRF KinEASE

Enzyme validation - Technical Notes

IRR, INSRR validated with HTRF KinEASE

Enzyme validation - Technical Notes

ITK validated with HTRF KinEASE

Enzyme validation - Technical Notes

JAK1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

JAK2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

JAK3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

KDR, VEGFR2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

KIT, c-kit validated with HTRF KinEASE

Enzyme validation - Technical Notes

LYNa validated with HTRF KinEASE

Enzyme validation - Technical Notes

MER,MERTK validated with HTRF KinEASE

Enzyme validation - Technical Notes

MET validated with HTRF KinEASE

Enzyme validation - Technical Notes

MUSK validated with HTRF KinEASE

Enzyme validation - Technical Notes

PDGFR-alpha, PDGFRA validated with HTRF KinEASE

Enzyme validation - Technical Notes

PDGFR-beta, PDGFRB validated with HTRF KinEASE

Enzyme validation - Technical Notes

PYK2, PTK2B validated with HTRF KinEASE

Enzyme validation - Technical Notes

RET validated with HTRF KinEASE

Enzyme validation - Technical Notes

RON, MST1R validated with HTRF KinEASE

Enzyme validation - Technical Notes

ROS, ROS1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

SRC validated with HTRF KinEASE

Enzyme validation - Technical Notes

SYK validated with HTRF KinEASE

Enzyme validation - Technical Notes

TIE2, TEK validated with HTRF KinEASE

Enzyme validation - Technical Notes

TRKA, NTRK1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

TRKB, NTRK2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

TRKC, NTRK3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

TYK2 validated with HTRF KinEASE

Enzyme validation - Technical Notes

TYRO3 validated with HTRF KinEASE

Enzyme validation - Technical Notes

YES1 validated with HTRF KinEASE

Enzyme validation - Technical Notes

ZAP70 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

Take advantage of HTRF to simplify HTS cascade

In collaboration with AstraZeneca - Posters

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 TK Kit / 62TK0PEB-62TK0PEC-62TK0PEJ

62TK0PEB-62TK0PEC-62TK0PEJ - 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

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