Discovery of GALNT3 Inhibitors Using a Universal Glycosyltransferase Assay and Orthogonal Pooled Screening

Transcreener UDP Assay Kit (FI) Method

Ordering Information

Instrument Compatibility

Related Resources

Transcreener UDP² Assay Kit - FI Readout

The Transcreener^® UDP² Assay Kit with FI readout directly measures UDP via antibodies selective to UDP and a fluorescent tracer. Enzymatically generated UDP displaces the tracer from an antibody-quencher conjugate, resulting in increased fluorescence. The universal detection method allows screening of virtually any glycosyltransferase in a high throughput format.

The Transcreener UDP Assay Kit with FI readout produces a simple fluorescent intensity output which can be read on even the most basic fluorescence readers typically found in academic and therapeutic research labs as well as more complex multimode plate readers. Direct detection of UDP with a fluorescence intensity (FI) readout provides a safe, HTS-compatible alternative to cumbersome radioassay methods and is more sensitive and less subject to interference than coupled assay methods, where the UDP is converted to another product.

Used in over millions of wells by pharma since 2006, Transcreener technology is the most extensively validated UDP detection method available backed by the best technical support in the industry.

This assay is designed to be used with purified enzyme preparations. Transcreener assays are not validated with cell lysates, blood, serum, or other biological samples.

Direct Detection of UDP to Measure Glycosyltransferase Enzymatic Activity

Fluorescence Intensity

Learn More About the Transcreener UDP Assay Kit

Transcreener UDP FI Assay Tech Manual

Applications

Measure enzymatic activity of glycosyltransferases
Screen compound libraries for glycosyltransferase modulators
Quantify Inhibitor Potency
Inhibitor Selectivity Profiling
Measure drug-target residence time

Features

Direct detection of unlabeled UDP
Easy to use, homogenous, one-step format
Robust Assay Z' > 0.7 under initial velocity conditions
Far-red fluorescent readouts minimize compound interference
A safe, non-radioactive method
Universal - use virtually with any glycosyltransferase

Easy-to-Use, Mix-and-Read, HTS Ready Assay

Run your enzyme reaction, add Transcreener reagents, and read your plates. The UDP assay kit is compatible with 96, 384. and 1536-well formats.

Example Applications

The Transcreener UDP Assay kit is compatible with any enzyme class that produces UDP, including UDP-glucose-, UDP-galactose-, & UDP-glucuronosyl-transferases-, as well as glycogen, hyaluranon, & cellulose synthetases.

Enzyme
Toxin B
GALNT2
GALNT3
BGalT1

Excellent Z' With Low Levels of UDP Conversion

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Far Red FI Readouts Validated on Major Multimode Readers

Supplier	Instrument	FI Assays
	TriStar²S LB 942	Validated
	Tristar 5	Validated
	Mithras² LB 943	Validated

	Cytation^TM 5	Validated
	Cytation^TM 3	Validated
	Cytation^TM 1	Validated
	Synergy^TM H1	Validated
	Synergy^TM 2/H4/4	Validated
	Synergy^TM HTX	Validated
	Synergy^TM Neo 2	Validated

	POLARstar^® Omega	Validated
	FLUOstar^® Omega	Validated
	PHERAstar^® FSX	Validated
	PHERAstar^® Plus/FS	Validated
	CLARIOstar^® /Plus	Validated
	VANTAstar^TM	Validated

	Sense	Validated

	Analyst^® GT/HT	Validated
	Gemini^® XPS/EM	Validated
	SpectraMax^® M2/M2e	Validated
	SpectraMax^® M5/M5e/FlexStation^® 3	Validated
	SpectraMax^® Paradigm	Contact Us
	SpectraMax^® iD3/iD5	Validated
	SpectraMax^® i3x	In Review

	EnVision^®/EnVision^® Xcite	Validated

	Infinite^® M1000/M1000Pro/Safire2^TM	Validated
	Infinite^® M200	Validated
	Infinite^® F500	Validated
	Infinite^® F200/Ultra^evolution	Validated
	Spark^TM 10M	Validated

Validation Criteria for Microplate Readers

384-Well Format
Z'-Factor ≥ 0.7 at 10% conversion of 10 μM ATP
Z' specifications to be met using Transcreener ADP² Assay Reagents
Read time to achieve Z' specifications in ≤ 5 minutes

Look for these validation stickers on instruments that have successfully met the validation criteria.

The instrument validation program is a collaborative effort with the major suppliers of multimode readers to insure that Transcreener assays perform optimally on your specific instrument. Scientists at BellBrook and/or the instrument vendor have tested Transcreener reagents with each instrument to determine optimal filters and settings for maximal assay performance. We provide you with application notes with all the important details, so you can be confident of success using Transcreener assays on the instrument of your choice.

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What's Included


Component	Notes
UDP² Antibody - IRDye^® QC-1	The concentration of antibody needed for an enzyme target is dependent upon the UDP concentration and buffer conditions in the enzyme reaction. Sufficient antibody is included in the kit to complete 200 assays (Part #3019-A), 1,000 assays (Part #3019-1K), or 10,000 assays (Part #3019-10K) at a UDP concentration up to 100 µM.
UDP Alexa Fluor^® 594 Tracer	The final tracer concentration in the 20 µL reaction is 4 nM.
Stop & Detect Buffer B, 10X	The Stop & Detect Buffer B components will stop enzyme reactions that require Mg²⁺. To ensure that the enzyme reaction is stopped completely, confirm that the EDTA concentration is at least equimolar to the magnesium ion concentration in the reaction. The final concentration of Stop & Detect Buffer B at the time of FI measurement is 0.5X.
UDP	UDP is used to create a donor substrate/UDP standard curve.

What You Will Need


Component	Notes
Glycosyltransferase Enzyme	Transcreener^® UDP assays are designed for use with purified enzyme preparations. Contaminating enzymes, such as phosphates or nucleotidases, can produce background signal and reduce the assay window.
Enzyme Buffer	User-supplied enzyme buffer components include enzyme, buffer, enzyme cofactors, substrates, and test compounds.
Assay Plates	Assay Plates Can Be Purchased Separately From BellBrook Labs. FI - An entirely black plate with a non-binding surface is required. We suggest Corning 384-well plates Cat. #4514.
Plate Reader	A multi-detection microplate reader configured to measure FI of the Alexa Fluor^® 594 tracer is required.
Liquid Handling Devices	Use liquid handling devices that can accurately dispense a minimum volume of 2.5 µL into 384-well plates.
Ultrapure Water	Some deionized water systems are contaminated with nucleases that can degrade both nucleotide substrates and products, reducing assay performance. Careful handling and use of ultrapure water eliminates this potential problem.

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Transcreener UDP² Assay Kits - FI Readout

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

Cat. No.	Product Description	Price	Add to Cart
3019-A	Transcreener UDP² FI Assay (200 Assay, 96 Well)	$359.00	Add to Cart
3019-1K	Transcreener UDP² FI Assay (1,000 Assay, 384 Well)	$759.00	Add to Cart
3019-10K	Transcreener UDP² FI Assay (10,000 Assay, 384 Well)	$4,299.00	Add to Cart
3019-100K	Transcreener UDP² FI Assay (10 X 10,000 Assay, 384 Well)	$21,495	Add to Cart

Assay Plates for FI

Cat. No.	Product Description	Price	Add to Cart
2233	Corning 96-Well Black Assay Plates (#3686), 3-Pack (200+ Assays)	$55.00	Add to Cart
2229	Corning 384-Well Black Assay Plates (#4514), 3-Pack (1,000+ Assays)	$60.00	Add to Cart
2230	Corning 384-Well Black Assay Plates (#4514), 30-Pack (10,000+ Assays)	$479.00	Add to Cart