Kinase Drug Discovery Services

Radiometric Kinase Activity Assays
Kinase Panel Screening
Cell-Based Kinase Assays
In Vivo Kinase Models
Recombinant Proteins
Custom Tailored Assay Services

Kinase Selectivity Assay Screening

Broad kinase screening is crucial for the identification of selective and potent kinase inhibitors. Determination and improvement of the selectivity of a compound by screening a large part of the kinome are of pivotal importance in the discovery process of novel kinase inhibitors e.g. to reduce the risk of adverse side effects or both, kinase inhibitors and compounds inhibiting non-kinase targets.

Reaction Biology offers the largest kinase panel for kinase profiling in the industry ensuring the most reliable results for your inhibitor selectivity screening covering most of the human kinome.
Kinase selectivity profiling is performed on a monthly (³³PanQinase™) or bi-weekly (HotSpot™) schedule, which allows us to offer very competitive pricing.
HotSpot kinase screening service now available at physiologically relevant 1mM ATP for 340 wild type kinases, in addition to existing standard concentrations of 1μM, 10μM or apparent ATP-Km up to 100μM
Specialty panels comprise a selection of kinase-specific mutants or our CDK panel.
Kinase panel reports from the US facility include a free control compound’s IC50 curve.

Kinase Selectivity Assay Panels

Panels run at US facility
Panels run at US facility	Panel size	ATP options	Testing options	Panel
Wild Type Kinase Panel	380	1μM, 10μM, Km and 1mM	1 & 10 doses	view panel
Diversify Panel	70	Km & 1mM	1 & 10 doses	view panel
Mutant Kinase Panel	348	1μM, 10μM, Km	1 & 10 doses	view panel
Atypical Kinase Panel	24	1μM, 10μM, Km	1 & 10 doses	view panel
Lipid Kinase Panel	23	1μM, 10μM, Km	1 & 10 doses	view panel
Diacylglycerol Kinase Panel	10	1μM, 10μM, Km	1 & 10 doses	view panel

Kinase Selectivity Assay Panels
Panels run at German facility	Panel size	ATP options	Testing options	Panel
Wild Type Kinase Panel	345	1μM, 10μM and Km	1 & 10 doses	view panel
Mutant Kinase Panel	96	1μM, 10μM, Km	1 & 10 doses	view panel
Lipid Kinase Panel	14	1μM, 10μM, Km	1 & 10 doses	view panel

Specialty Panels

Panels run at US facility	Panel size	Data
ABL1 Mutant Profiler	13	Data sheet
ALK Mutant Profiler	28	Data sheet
BRAF Mutant Profiler	16	Data sheet
c-KIT Mutant Profiler	22	Data sheet
c-MET Mutant Profiler	30	Data sheet
CDK Profiler	34	Data sheet
EGFR Mutant Profiler	60	Data sheet
ERBB2 Mutant Profiler	9	Data sheet
FGFR2 Mutant Profiler	13	Data sheet
FGFR3 Mutant Profiler	6	Data sheet
FGFR4 Mutant Profiler	5	Data sheet
FLT3 Mutant Profiler	9	Data sheet
RET Mutant Profiler	34	Data sheet
TIE Mutant Profiler	10	Data sheet
TRKA Mutant Profiler	15	Data sheet
TRKC Mutant Profiler	6	Data sheet

Panels run at German facility	Panel size	Data
ABL1 Mutant Profiler	9	Data sheet
ALK Mutant Profiler	8	Data sheet
CDK Profiler	32	Data sheet
EGFR Mutant Profiler	11	Data sheet
KIT Mutant Profiler	10	Data sheet
Lipid Kinase Profiler	14	Data sheet
MET Mutant Profiler	10	Data sheet
RET Mutant Profiler	10	Data sheet

Kinase Assay Details

Assay Formats

HotSpot™ and ³³PanQinase™: two radiometric assay formats

Principle of radiometric kinase activity assay for kinase inhibitor screening.. Radioactive phosphate get's transferred to the kinase substrate. the now radioactive substrate get's quantified.

Reaction Biology offers two radiometric assay formats that are performed in our screening facilities located in Malvern, PA, USA, and Freiburg, Germany. Both assays are based on the transfer of ³³P-labelled phosphate from ATP to the kinase substrate. The methods, however, differ with respect to retention and detection of the phosphorylated substrates. Proteins and peptides in the HotSpot™ assay (performed in the US) are captured via spotting of the reaction mix on a filter membrane. The ³³PanQinase™ assay (performed in Germany) is performed with ScintiPlate microtiter plates, which are coated with scintillant for detection.

Lipid kinases are screened with ADP-Glo (Promega) at both facilities.

Assay Advantages

Radiometric assay formats are the gold standard for kinase screening producing reproducible, high-quality data, and directly measure enzyme activity. The catalytic product is directly detected (phosphorylated peptide or protein). False positives and negatives, as caused by indirect formats of detection, are avoided.

Both, the HotSpot^TM and 33PanQinase^TM assays allow detection of ATP- as well as substrate competitive and allosteric inhibitors.

HotSpot^TM assays are adaptable to any protein or peptide substrate without modification.

Download a comprehensive overview on “Kinase Profiling & Screening- Choosing a Biochemical Assay Platform”

Screening at US Facility

Setups: Single-dose screening in duplicates or IC50 value determination with 5 or 10 concentrations. Other screening formats are available upon request.

The assay protocol for HotSpot™ Assay is available here.

ATP concentrations: 1mM, 1μM, 10μM, or apparent ATP-Km up to 100μM.

Controls: No inhibitor control (DMSO vehicle) and for every kinase, one kinase-specific control compound is tested in 10-dose IC50 format.

Screening facility: Malvern, PA, USA

Turnaround time: 10 business days for standard projects. Expedited scheduling and data delivery can be arranged prior to the commencement of the studies.

Report: The raw data, % enzyme activity and control compound IC50 values will be reported in Excel format for single-dose assays. For IC50 orders, raw data, IC50 values, and curve fitting will be delivered in Excel format.
Assay conditions, target, and substrate information are available upon request. Requirements for this information should be noted prior to the commencement of the study.

Compound requirements: In brief, for a standard project with less than 50 kinases, 20 µl of a 10 mM DMSO stock or solid material is needed. Please inquire for compound requirements for large scale screening. Please refer to our FAQs for information regarding compound preparation and shipping.

Please download our assay condition form (US facility).

Screening at German Facility

Setup: Single-dose screening or IC50 value determination with 10 concentrations. Other screening formats are available upon request.

The assay protocol for ³³PanQinase™ assay is available here.

ATP concentrations: apparent ATP-Km

Controls: DMSO only control; reference inhibitor control for IC50 value measurement

Screening facility: Freiburg, Germany

Turnaround time: 10 business days for standard projects. Expedited scheduling and data delivery can be arranged prior to commencement of the studies

Report: A detailed report will be provided including assay conditions, target, and substrate information. The raw data and % enzyme activity will be reported in Excel including IC50 values and curve fitting, if applicable.

Compound requirements: In brief, for a standard project with less than 33 kinases, 100 µl of the stock solution with 100x of the highest assay concentration or solid material. Please inquire for compound requirements for large scale screening.

Please download our assay condition form (German facility) for further details on compound preparation and shipping.

High-Throughput Screening

Library screening: Screen your own or our libraries.

Available libraries:

Drug-like diversified small molecule libraries; > 110,000 compounds
Focused small libraries including kinase inhibitors, FDA-approved inhibitors, epigenetic inhibitors, natural products, and a clinical collection.
Fragment libraries including a standard diversified fragment library, Fsp3-enriched fragments library, 3D fragment library, protein-protein interaction fragments, and covalent fragments library.

In addition to our high-throughput primary screening formats, we offer orthogonal screening formats and secondary screening formats. Hits can be characterized for their kinetic, binding affinity, mechanism of action, and intracellular activity.

Our medicinal chemistry partners are available to optimize the structure-activity-relationship of your hits to increase potency and reduce off-target and toxic effects and enhance the pharmacological profile of your lead compounds.

Cellular Kinase Assay Screening

Cell-based assays are a great choice for secondary screening after initial hit finding because of their inherent nature of testing compound action in intact, living cells.

To facilitate science-driven kinase drug discovery, Reaction Biology has created three different cellular platforms for kinase inhibitor screening in intact cells.

How do they compare?

Assay	NanoBRET Target Engagement Assay	Cellular Phosphorylation Assay	BaF3 Cell Proliferation Assay
Readout	Compound binding to the kinase	Activity of the kinase	Cell transformation capability of the kinase
Parameters	Binding affinity, target residence times, target occupancy, binding kinetics	Inhibitor potency	Inhibitor potency, kinase-engaged signaling pathways, transformation capacity of the kinase
Assay type	Competitive tracer displacement	Substrate phosphorylation	Cell survival and proliferation
Assay technology	BRET: energy transfer caused by compound-kinase proximity resulting in luminescence	ELISA or AlphaLISA to quantify phosphorylated substrate	Cell proliferation assay via Cell Titer Glo
Detectable inhibitor types	All	All	All
Kinase families	All	Protein kinases	Oncogenic ‘driver’ kinases
Kinase expression	Exogenous	Endogenous or exogenous	Exogenous
Compound incubation period	1-2 hours	1-2 hours	2-3 days

Our blog Spotlight: Cell-based kinase assay formats. describes our three assay formats in more detail. Check it out!

Why Choose Reaction Biology for Kinase Assay Research

We’ll help you develop an optimal treatment for cancer and other diseases. For 20 years, we’ve accumulated the largest portfolio of kinase assays available. You can trust our expertise in providing tailored assay development. Work with us and you’re guaranteed to get the exact in vitro kinase assay you need.

Why Choose Reaction Biology