Reliable, integrated tools for drug discovery

GeneBLAzer® beta-lactamase Reporter Technology provides you with the most reliable, integrated set of tools for accelerating your cell-based target validation, pathway analysis, and compound screening in drug discovery.

This proven technology combines molecular and cell biology and a Fluorescence Resonance Energy Transfer (FRET)-based, ratiometric detection method that reduces experimental noise which can lead to false hits.

The GeneBLAzer® Technology consists of readily accessible tools and custom assays (click table below) for studying numerous target classes and cellular processes, including: surface and intracellular reporters (such as nuclear and cytokine receptors, orphan and known G-protein coupled receptors), a wide range of signal transduction pathways, ion channels, other transporters, and transcriptional regulators (including kinases and intracellular processes).

Integrated Solutions Using GeneBLAzer® Reporter Technology

GeneBLAzer Reporter Technology

How GeneBLAzer® Technology works

GeneBLAzer® Technology uses a mammalian-optimized bla gene combined with a FRET-enabled substrate to provide reliable and sensitive detection in cells.

Cells are loaded with an engineered fluorescent substrate containing two fluoroprobes, coumarin and fluorescein. In the absence of bla expression, the substrate molecule remains intact. In this state, excitation of the coumarin results in fluorescence resonance energy transfer to the fluorescein moitey and emission of green light. However, in the presence of bla expression, the substrate is cleaved, separating the fluorophores, and disrupting energy transfer. Excitation of the coumarin in the presence of enzyme bla activity results in a blue fluorescence signal. The resulting blue:green ratio provides a normalized reporter response.

GeneBLAzer FRET Enabled Substrate


Ratiometric Advantage of GeneBLAzer® Technology: Minimize the Effects of Experimental Noise

To compare the effects of some types of experimental noise on ratiometric and non-ratiometric reporter readouts, data were generated using assay conditions that included variations in cell number and substrate concentration. Cells constitutively expressing beta-lactamase were plated in varying degrees of confluency and LiveBLAzer™ - FRET B/G Substrate was added in either the recommended (1X) or high (4X) concentration.
Ratiometric Non Ratiometric Responses
Data for the ten samples was then analyzed using either the blue/green (ratiometric) or blue (non-ratiometric) values from this assay. The data illustrates the ratiometric readout reduces both absolute and relative errors caused by experimental noise compared to non-ratiometric methods.

Full, Partial, and Inverse Agonists Distinguished using GeneBLAzer® Technology

HEK 293T cells expressing beta-lactamase coupled to a cyclic AMP response element (CRE) were stably transfected with a cDNA encoding a G-Protein Coupled Receptor. Cells were then treated with increasing concentrations of several agonists and analyzed using LiveBlazer™ - FRET B/G Substrate. The data shows the ability of the GeneBLAzer® Technology to establish rank order of potency and to distinguish types of agonism, even when the differences between them are small. Dose response curves shown are full agonists (Compounds 1 and 2), partial agonists (Compounds 3 and 4) and an inverse agonist (Compound 5).

Multiplexing GeneBLAzer® Technology with Fluo-4

5HT1a-G_NFAT-bla CHO-K1 cells were run in agonist mode with the LOPAC 12080TM library of small molecules using Fluo-4 multiplexed with the LiveBLAzer™-FRET B/G Assay. Cells were loaded with Fluo-4 to search for compounds that trigger the Ca2+ flux upon binding to the serotonin receptor. The same cells were loaded with LiveBLAzer™-FRET B/G substrate and then analyzed. Hits registered with Fluo-4 are shown in Panel 1, and hits registered with LiveBLAzer™-FRET B/G are shown in Panel 2.

Multiplexing GeneBLAzer Fluo-4Out of 18 known serotonin receptor agonists detected by the LiveBLAzer™ Assay, 16 were also detected by the Fluo-4 Assay. In contrast, 4 other compounds detected by LiveBLAzer™ Assay were not detected by the Fluo-4 Assay, while 6 additional compounds detected by the Fluo-4 Assay were not detected by the LiveBLAzer™ Assay. By multiplexing these two assay technologies, one can validate true hits, while eliminating those that are not.