Profiling PDEs with the Transcreener AMP²/GMP² Phosphodiesterase Assay

Transcreener AMP²/GMP² Phosphodiesterase Assay Kit

Detect any AMP or GMP producing enzyme (e.g., ligases, synthetases, phosphodiesterases) using any precursor substrate, including cAMP, cGMP, ATP, or NAD. It is the only activity assay method for direct detection of unlabeled AMP or GMP; i.e. without using coupling enzymes. The method is simple and HTS-proven: run your enzyme reaction, add detection reagents, and read the far red FP or TR-FRET signal on any multimode plate reader.

XMP enzyme products, including AMP, GMP, and CMP displace a fluorescent tracer from a highly selective antibody resulting in a change in fluorescence. Both assays use a far-red tracer, which reduces compound interference and provides a robust ratiometric readout. Make measuring cyclic nucleotide phosphodiesterase activity easy with a simple, proven, HTS-ready assay.

Universal AMP/GMP Detection for Phosphodiesterases

FP AMP/GMP Assay - LEARN MORE

Phosphodiesterase Assay Kit Schematic AMP GMP Assay FP

Transcreener AMP²/GMP² Assay Tech Manual – Fluorescence Polarization (PDF)

TR-FRET AMP/GMP Assay - LEARN MORE

Phosphodiesterase Assay Kit Schematic AMP GMP Assay TR-FRET

Transcreener AMP²/GMP² Assay Tech Manual – TR-FRET (PDF)

One Assay … Hundreds of Targets

Along with PDEs the kit can also be used with:

Ligases: Ub and SUMO Ligases, DNA and RNA Ligases
Synthetases: Acyl CoA Synthetase, Amino-Acyl tRNA Synthetases, NAD Synthetase
Phosphodiesterases: cAMP, cGMP
Sialyltransferases: (CMP detection)
Protein deAMPylase
Ectonucleotidases such as CD39, CD73, and ENPP1

Applications

Measure Enzymatic Activity
Screen Compound Libraries for Inhibitors
Determine Inhibitor Selectivity
Measure Inhibitor Potency
Inhibitor Residence Time Measurement

Measure the Potency of Inhibitors for a Variety of Targets

Features

Direct detection of unlabeled AMP or GMP from a PDE reaction
Easy to use: one-step, simple mix-and-read format
Robust: Z’ > 0.7 for initial velocity reactions
Less interference: Far-red fluorescence minimizes compound interference
Non-radioactive assay technique
Available in FP or TR-FRET readouts
Wide substrate concentration range: 1 μM to 1000 µM

Sensitive Assay with Large Window

PDE Assay Services

Interested in moving your program forward, but don't want to bring an assay in-house? Our scientists can help! BellBrook scientists will use their extensive biochemistry and enzymology expertise to work with you and accelerate your PDE program.

Lead Discovery Services Include:

Inhibitor Screening – To identify or confirm activity with the target.
Inhibitor Potency Profiling – Dose-response with target and/or related proteins. Fast IC₅₀ results.
Residence Time Measurements – Determination of k_off using ‘jump dilution’ enzymatic assay method.
Mechanism of Action Studies – Kinetic analysis to define the mode of inhibition.

Contact Us to Learn More

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What's Included

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Component	Notes
AMP/GMP Antibody	Antibody specific to the nucleotides AMP and GMP.
AMP/GMP Tracer	When displaced, changes in fluorescence can be read in an FP or TR-FRET depending on the assay format.
Tris Solution	Tris solution is used to buffer the detection mixture.
AMP	AMP is used to create the cAMP/AMP standard curve.
GMP	GMP is used to create the cGMP/GMP standard curve.

What You Will Need

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Component	Notes
Enzyme	The assays are designed for use with purified enzymes. Use any enzyme that produces AMP, GMP, or CMP.
Enzyme Buffer	Enzyme buffer components optimized for your target. See the technical manual for potential interfering components.
ATP, cAMP, cGMP	Use the nucleotide required for your enzyme. It is possible to purchase some nucleotides direct from BellBrook Labs.
Assay Plates	Assay Plates Can Be Purchased Directly Through BellBrook Labs FP - An entirely black plate with a non-binding surface is required. We suggest Corning 384-well plates Cat. #4514 TR-FRET - An entirely white plate with a non-binding surface is required. We suggest Corning 384-well plates Cat. #4513
Stop Buffer	Some PDEs and Ubiquitin Ligases have inhibited activity with Stop Buffer B (Cat. #2027-1 mL, Cat. #2032 10 mL) which quenches MgCl₂. This allows the assay to be run in endpoint mode.

Far Red FP & TR-FRET Readouts Validated on Major Multimode Readers

Instrument	FP Assays	TR-FRET Assays
TriStar²S LB 942	in review	validated
Mithras² LB 943	in review	validated

Cytation™ 5	validated	validated
Cytation™ 3	validated	validated
Cytation™ 1	validated	validated
Synergy™ H1	validated	validated
Synergy™ 2/H4/4	validated	validated
Synergy™ HTX	not capable	not capable
Synergy™ Neo 2	validated	validated

POLARstar^® Omega	validated	validated
FLUOstar^® Omega	not capable	validated
PHERAstar^® FSX	validated	validated
PHERAstar^® Plus/FS	validated	validated
CLARIOstar^® /Plus	validated	validated

Analyst^® GT/HT	validated	validated
Gemini^® XPS/EM	not capable	not capable
SpectraMax^® M2/M2e	not capable	not capable
SpectraMax^® M5/M5e/FlexStation^® 3	not capable	validated
SpectraMax^® Paradigm	validated	contact us
SpectraMax^® iD3/iD5	in review	validated

EnVision^®/EnVision^® Xcite	validated	validated

Infinite^® M1000/M1000Pro/Safire2™	validated	validated
Infinite^® M200	not capable	not capable
Infinite^® F500	validated	validated
Infinite^® F200/Ultra^evolution	contact us	contact us
Spark™ 10M	validated	validated

Transcreener AMP²/GMP² Phosphodiesterase Assay Kits

* For custom or bulk orders (over 100,000 wells) please contact us (info@bellbrooklabs.com) for a quote.

Edit
Cat. No.	Product Description	Price	Add to Cart
3015-1K	Transcreener AMP²/GMP² FP Assay (1,000 Assay, 384 Well)	$639.00	Add to Cart
3015-10k	Transcreener AMP²/GMP² FP Assay (10,000 Assay, 384 Well)	$3,689.00	Add to Cart
3015-100k	Transcreener AMP²/GMP² FP Assay (10 X 10,000 Assay, 384 Well)	$18,445.00	Add to Cart
3020-1K	Transcreener AMP²/GMP² TR-FRET Assay (1,000 Assay, 384 Well)	$699.00	Add to Cart
3020-10K	Transcreener AMP²/GMP² TR-FRET Assay (10,000 Assay, 384 Well)	$3,989.00	Add to Cart
3020-100K	Transcreener AMP²/GMP² TR-FRET Assay (10 X 10,000 Assay, 384 Well)	$19,945.00	Add to Cart

Assay Plates for FP

Cat. No.	Product Description	Price
2233	Corning 96-Well Black Assay Plates (#3686), 3-Pack (200+ Assays)	$40.00	Add to Cart
2229	Corning 384-Well Black Assay Plates (#4514), 3-Pack (1,000+ Assays)	$45.00	Add to Cart
2230	Corning 384-Well Black Assay Plates (#4514), 30-Pack (10,000+ Assays)	$400.00	Add to Cart

Assay Plates for TR-FRET

Cat. No.	Product Description	Price
2234	Corning 96-Well White Assay Plates (#3642), 3-Pack (200+ Assays)	$40.00	Add to Cart
2231	Corning 384-Well White Assay Plates (#4513), 3-Pack (1,000+ Assays)	$50.00	Add to Cart
2232	Corning 384-Well White Assay Plates (#4513), 30-Pack (10,000+ Assays)	$450.00	Add to Cart

PDE's As Enzyme Drug Targets

Cyclic nucleotide phosphodiesterases are proven drug targets. Most are a focus of inhibition due to their functional properties. Since PDEs degrade cAMP or cGMP, inhibitors can prolong the effects of the cyclic nucleotide in the signaling cascade within the tissues where the individual enzyme is most likely to reside.

PDEs are currently classified into 11 different families. Each is classed based on a number of different factors including the distribution within tissue, and substrate specificity. Some hydrolyze cAMP, while others cGMP. A variety can degrade both cAMP and cGMP. The Transcreener antibody can detect the product AMP or GMP of cyclic nucleotide hydrolysis. This allows the measurement of virtually any PDE's activity.

Substrate Specificity

Nucleotide	PDE
cAMP	4, 7, 8
cGMP	5, 6, 9
cAMP & cGMP	1, 2, 3, 10, 11

The Transcreener phosphodiesterase assay enables researchers to scour vast compound libraries for potent, yet selective inhibitors for unique PDEs. Follow-up SAR can also be completed to improve upon initial hits and provide a greater probability for success. PDEs have excellent promise in a variety of therapeutic areas including heart disease, dementia, infection, depression, and asthma. New inhibitors can provide better therapeutic potential and can limit off-target effects more effectively.

Transcreener® AMP²/GMP² Phosphodiesterase Assay

Profiling PDEs with the Transcreener AMP²/GMP² Phosphodiesterase Assay

Transcreener AMP²/GMP² Phosphodiesterase Assay Kit

Universal AMP/GMP Detection for Phosphodiesterases

Transcreener AMP²/GMP² Assay Tech Manual – Fluorescence Polarization (PDF)