BellBrook Labs Transcreener AMP/GMP Assay

BellBrook Labs Transcreener™ AMP/GMP Assay is a far-red, competitive fluorescence polarization assay based on the detection of AMP or GMP, and is therefore compatible with any enzyme class that produces either AMP or GMP, including ubiquitin, SUMO, nucleic acid and protein ligases, phosphodiesterases (PDEs), and synthetases. The Bell Brook Lab's Transcreener AMP/GMP Assay is a single step, homogenous detection assay enabling the use of unmodified native substrate concentrations of 1 - 1000 μM. The assay provides excellent signal at low substrate conversion, with a Z’ ≥ 0.7 and ≥ 85 polarization shift (mP) at 10% substrate conversion using 10 μM donor substrate.


Benefits

•  Single-step reaction using unlabeled, native cAMP, cGMP and ATP substrates
•  No "in-the-dark" steps. No manual steps. No secret reagents.
•  Wide substrate concentration range [ 1 μM to 1000 μM]
•  Real-time or endpoint detection

Description

Phosphodiesterases control a variety of cellular processes by hydrolyzing the second messenger signaling molecules cAMP and cGMP. The Transcreener AMP/GMP Assay is a far-red, competitive fluorescence polarization immunoassay that detects the reaction products AMP or GMP. Enzyme reaction progress is indicated by a decrease in the fluorescence polarization signal. The Transcreener AMP/GMP Assay is a simple two-step, endpoint assay accomodating cAMP/cGMP concentrations of 0.1 to 10 μM with a single reagent mix (up to 1 mM substrate has been used). The assay provides an excellent signal under initial velocity conditions resulting in overall Z’> 0.6. Although originally designed for PDEs that hydrolyze cAMP or cGMP, the Transcreener AMP/GMP Assay can be used for any enzyme class that produces either AMP or GMP (eg: a ligase reaction which converts ATP to AMP). Reduced development costs and accelerated drug discovery are achieved by utilizing this one simple streamlined screening method.

The Transcreener AMP/GMP Detection Mixture comprises an AMP/GMP Alexa633 tracer bound to an AMP/GMP antibody, which is displaced by AMP or GMP, the products generated during PDE reactions. The displaced tracer freely rotates leading to a decrease in fluorescence polarization, relative to the bound tracer. Therefore, AMP/GMP production creates a proportional decrease in polarization values.