Cell-Based In Vitro Kinase Assay Services

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.

Compounds face multiple challenges when put to work in cells such as

they need to cross the plasma membrane when targeting intracellular kinases
they meet an abundance of other proteins that they may bind to than the target kinase, potentially leading to off-target effects causing adverse drug reactions
they are faced with the physiological pH, co-factors, ion concentrations in cells
they must act on the target in its tertiary structure complete will all post-translational modifications such as glycosylations

Read more about the importance of screening compounds in cell-based assays in our blog: Testing kinase inhibitors where it matters: Drug screening in intact 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!

BaF3 Cell Proliferation Assay

The BaF3 Cell Proliferation Assay is an activity assay for the investigation of the potency of inhibitors suppressing oncogenic kinases. It is useful for the investigation of kinase-engaged signaling pathways and testing of off-target effects of kinase inhibitors.

Cellular Phosphorylation Assay

The Cellular Phosphorylation Assay is a kinase activity assay performed in intact, living cells. The assays have been designed to address compound activity in a physiological environment measuring changes in the phosphorylation status of physiological substrates.

NanoBRET TE Intracellular Kinase Assay

The NanoBRET Intracellular Kinase Assay measures the binding of a compound to the target kinase in intact, living cells. Allowing assessment of target occupancy, binding affinity, and kinetics this assay allows insight into the compound-target interaction in the physiologic environment of cells.