Transcreener® HTS Assays
Four Assays Cover Thousands of Target Enzymes
Universal, high throughput screening platform for enzymes based on direct detection of nucleotide enzyme products with far red FI, FP and TR-FRET readouts.
- Four assays cover thousands of target enzymes, including any kinase, ATPase, or GTPase
- Direct detection means less chance for compound interference
- Far Red FP, FI and TR-FRET readouts validated on major multimode readers
- Single step, mix and read format with 8 hour deck and signal stability for easy automation
- Transcreener assays enable you to screen more targets, both well characterized and emerging, faster and more efficiently. Thousands of cellular reactions use nucleotides, either alone or in combination with other substrates. Enzymes in the purinome alone - those that use adenine and guanine nucleotides - comprise 13% of the coding capacity of the human genome1. Of these, protein kinases have become highly validated drug targets, but lipid kinases, chaperone ATPases and GTPases as well as less obvious ATP-dependent enzymes such as Acetyl Co-A carboxylase are increasingly being targeted for therapeutic intervention.
Bellbrook Labs' assays cover thousands of target enzymes, including any kinase, ATPase, or GTPase
- Transcreener is a universal assay method that can be used across entire families of nucleotide-dependent enzymes. Rather than using separate assays for a multitude of specific reaction products, such as phosphorylated proteins or lipids, a single nucleotide detection assay can be used for all of the enzymes that generate a common nucleotide product.
Transcreener Assay using the GDP FI Assay as an example. Displacement of the tracer from antibody by GDP relieves quenching resulting in increased fluorescence.
- Direct detection of nucleotides is unique to the Transcreener platform. The method is based on the interaction of two detection reagents: an antibody and tracer. Displacement of tracer by the nucleotide being detected causes a change in its fluorescence properties. In the case of the TR-FRET and FI formats, the antibody is covalently labeled, whereas for FP it is unmodified.
- Transcreener assays have fewer reagents and a less complex mechanism than any other nucleotide detection assay, all of which require coupling enzymes. Coupling enzymes are used to convert a nucleotide to a product that can be used by a reporter enzyme to generate a signal. Each coupling and reporter enzyme is a potential target for the compounds being screened, which increases the risk of false positives or of missing a hit.
- The Transcreener platform relies on highly specific antibodies that are able to differentiate between nucleotides on the basis of a single phosphate or even a methyl group. Selectivities for the product nucleotides vs. the substrates (e.g., ADP vs. ATP) range from 150-fold to over 1000-fold. Differentiation between closely related nucleotides is central to the technology as it allows detection of nucleotide enzyme products in the presence of an excess of the substrate nucleotide (e.g, ADP detection in the presence of excess ATP), a requirement for measuring enzyme initial velocity.
- All of the Transcreener assays generate Z' values of greater than 0.7 at substrate conversion levels of less than 10% - usually far lower – over the full range of initial substrate concentration.
PKA titration using 20µM ATP showing sensitive detection of very low levels of ADP formation.
BellBrook's partners in the Transcreener Instrument Validation Program.
- In addition to giving you flexibility of three detection modes, BellBrook has collaborated with the major suppliers of multimode readers to optimize instrument hardware and software settings for maximal performance with each of the Transcreener Assays. Successful validation requires a Z' of greater than 0.7 for nucleotide detection under initial velocity conditions (≤10% conversion). Optimal filter sets and instrument settings are summarized in the Technical Manual for each Transcreener assay.This ensures that whatever fluorescent detection mode or reader you prefer, you will get robust results with Transcreener assay reagents.
Transcreener assays are homogenous, single addition methods.
- Direct detection also means that the protocol is the simplest available: run your enzyme reaction, add Transcreener reagents with stop mix, and read plates. Some coupled methods require extra liquid addition and incubation steps, which complicates assay automation.
- The Transcreener antibodies and tracers are very stable molecules, which allow the detection mix to be pre-mixed and stored for long periods at room temperature. Also, unlike the transient signals from some coupled enzyme assays, the Transcreener fluorescent signals persist at least overnight – some for days - so plates can be read long after the detection reagents are added. These properties make Transcreener very easy to use in an automated HTS environment.
The Most Highly Validated ADP Detection Method -Transcreener® ADP2 Assay-Fluorescence Polarization
- Transcreener ADP2 FP Assay uses a highly sensitive antibody against ADP and a far red tracer to produce an excellent signal at less than 10% ATP consumption for a broad range of initial ATP concentrations (0.1-1,000 μM). Both reagents and signal are stable at room temperature for over 24 hours providing outstanding flexibility for automated high-volume screening programs. Used in over 40M wells of pharma screening since 2006, Transcreener is the most extensively validated ADP detection method available backed by the best technical support in the industry.
Principle of ADP2 Fluorescence Polarization Assay
- The Transcreener ADP2 FP Assay measures the progress of any enzyme that produces ADP. Displacement of the tracer by ADP causes a decrease in fluorescence polarization. The assay uses a red-shifted tracer, which reduces compound interference and provides a robustratiometric readout.
Transcreener® Performance with a Positive Readout - Transcreener® ADP2 Assay-Fluorescence Intensity
- The Transcreener® ADP2 Fluorescent Intensity (FI) Assay 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. It is a simple one-step homogenous detection assay, and is flexible with regard to ATP concentration (0.1 to 1,000 µM ATP). The assay provides excellent signal at low substrate conversion, with a Z' ≥ 0.7 at 2.5% ATP conversion using 1 µM ATP.
Displacement of tracer by ADP relieves quenching resulting in a positive fluorescence intensity signal.
Universal ADP Detection with a Far-Red TR-FRET Readout - Transcreener® ADP2 Assay-TR-FRET
- The Transcreener® ADP2 TR-FRET Red Assay produces a far-red, time-resolved Förster-resonance-energy-transfer (TR-FRET) signal that is less sensitive to interference from fluorescent compounds than other detection modes. It is a simple, one-step homogenous detection assay that accommodates a broad range of ATP concentrations (0.1 to 1,000 µM ATP) and produces excellent signal (Z' ≥ 0.7 at 10% conversion of 1 µM ATP).
Displacement of tracer by ADP disrupts FRET from the antibody-bound Terbium chelate leading to a decrease in the TR-FRET signal. In addition to the time-resolved signal, the assay uses a far-red tracer to minimize interference from fluorescent compounds and light scattering.
Universal AMP2/GMP2 detection – Transcreener® AMP2/GMP2 Assay
- The Transcreener AMP2/GMP2 Assay allows you to detect any AMP2/GMP2 producing enzyme (e.g., ligases, synthetases, phosphodiesterases) using any precursor substrate, including cAMP, cGMP, ATP, or NAD. The method is simple and HTS-proven: run your enzyme reaction, add detection reagents, and read the far red FP 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 decrease in fluorescence polarization. The assay uses a far red tracer, which reduces compound interference and provides a robustratiometric readout.
The only Mix-and-Read Fluorescent GTPase Assay
- The Transcreener GDP Assays are single step, homogenous, fluorescent assays for direct detection and screening of GTPases. Available with fluorescence polarization (FP), fluorescence intensity (FI), and TR-FRET readouts, all far red, they provide a safe, HTS-compatible alternative to cumbersome radioassay methods and are more sensitive and less subject to interference than colorimetric phosphate detection methods.
- Based on BellBrook's proprietary nucleotide immmunodetection technology, the Transcreener GDP Assays are compatible with any enzyme class that produces GDP, including monomeric small G proteins, such as CDC42, and Ga subunits of heterotrimeric G proteins, such as Gai1.
Principle of Fluorescence Polarization Assay
Principle of Fluorescence Intensitiy Assay
- GDP displaces tracer from a highly selective monoclonal antibody resulting in a decrease in fluorescence polarization or an increase in fluorescence intensity.
Universal UDP-Glycosyltransferase Assays
- Glycosylation controls the function and localization of thousands of human proteins and is central in bacterial cell wall synthesis. Selective targeting of glycosyltransferases may offer new avenues for therapeutic intervention in diverse diseases, including cancer, inherited metabolic disorders, and infectious diseases. Transcreener is the only HTS-compatible glycosyltransferase assay method that relies on direct detection of reaction products and does not depend on coupling enzymes, which are prone to interference.
Transcreener® UDP2 Assay – Fluorescence Polarization
- The Transcreener UDP2 Assay is a single step, homogenous, fluorescent assay for detection and screening of UDP-producing enzymes. Direct detection of UDP with a fluorescence polarization (FP) 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. The Transcreener UDP2 Assay is compatible with any enzyme class that produces UDP, including UDP-glucose-, UDP-galactose- and UDP-glucuronosyl-transferases as well as glycogen, hyaluranon and cellulose synthases.
New! Transcreener® UDP2 Assay – TR-FRET
- Homogenous detection of UDP producing enzymes (including UDP-glucose-, UDP-galactose-, and UDP-glucuronosyl-transferases as well as glycogen, hyaluranon, and cellulose synthases) is now possible with a TR-FRET readout. Like the original, FP-based assay the Transcreener UDP2 TR-FRET assay is designed for automated HTS platforms: run your enzyme reaction, add detection reagents, and read the far red TR-FRET signal on any multimode plate reader using the same filter sets as HTRF®. The TR-FRET detection mode relies on a time-gated luminescence signal, making it resistant to interference from fluorescent compounds in chemical libraries.
Principle of Fluorescence Polarization Assay
UDP2 Assay with TR-FRET Readout
- For shipping and storage information please click on Order#.
|3010-1K||Transcreener ADP2 FP Assay||EA||567,00|
|3010-10K||Transcreener ADP2 FP Assay||EA||3117,00|
|3010-100K||Transcreener ADP2 FP Assay||EA||on request|
|3011-1K||Transcreener ADP2 TR-FRET Red Assay||EA||654,00|
|3011-10K||Transcreener ADP2 TR-FRET Red Assay||EA||3927,00|
|3011-100K||Transcreener ADP2 TR-FRET Red Assay||EA||on request|
|3013-A||Transcreener ADP2 FI Assay||EA||282,00|
|3013-1K||Transcreener ADP2 FI Assay||EA||567,00|
|3013-10K||Transcreener ADP2 FI Assay||EA||3401,00|
|3013-100K||Transcreener ADP2 FI Assay||EA||on request|
|3015-1K||Transcreener AMP2/GMP2 FP Assay||EA||567,00|
|3015-10K||Transcreener AMP2/GMP2 FP Assay||EA||3401,00|
|3015-100K||Transcreener AMP2/GMP2 FP Assay||EA||on request|
|3020-1K||Transcreener AMP2/GMP2 TR-FRET Assay||EA||654,00|
|3020-10K||Transcreener AMP2/GMP2 TR-FRET Assay||EA||3927,00|
|3020-100K||Transcreener AMP2/GMP2 TR-FRET Assay||EA||on request|
|3009-1K||Transcreener GDP FP Assay||EA||567,00|
|3009-10K||Transcreener GDP FP Assay||EA||3401,00|
|3009-100K||Transcreener GDP FP Assay||EA||on request|
|3021-1K||Transcreener GDP TR-FRET Assay||EA||654,00|
|3021-10K||Transcreener GDP TR-FRET Assay||EA||3927,00|
|3021-100K||Transcreener GDP TR-FRET Assay||EA||on request|
|3014-A||Transcreener GDP FI Assay||EA||282,00|
|3014-1K||Transcreener GDP FI Assay||EA||567,00|
|3014-10K||Transcreener GDP FI Assay||EA||3401,00|
|3014-100K||Transcreener GDP FI Assay||EA||on request|
|3018-1K||Transcreener UDP2 FP Far Red Assay||EA||567,00|
|3018-10K||Transcreener UDP2 FP Far Red Assay||EA||3401,00|
|3018-100K||Transcreener UDP2 FP Far Red Assay||EA||on request|
|3022-1K||Transcreener UDP2 TR-FRET Assay||EA||654,00|
|3022-10K||Transcreener UDP2 TR-FRET Assay||EA||3927,00|
|3022-100K||Transcreener UDP2 TR-FRET Assay||EA||on request|
- All prices are in EURO excl. VAT and shipping. Only available in selected countries.
Downloads, Links and Webinars
- Transcreener® HTS Assays Brochure (PDF)
- Probes for Investigating Signal Transduction (Brochure) (PDF)
- Transcreener® ADP2 Assay Flyer (PDF)
- Transcreener® AMP2/GMP2 Assay Flyer (PDF)
- Transcreener® FAQ’s (Link)
- Transcreener® Posters (Link)
- Transcreener® Instrument Validation (Link)
- Transcreener® Publications (Link) on www.bellbrooklabs.com
- Webinar: Transcreener® HTS Assays for UDP, GDP, and CMP as a Universal Glycosyltransferase Assay Platform View Now
- Webinar: Analyzing Kinase Inhibitor Residence Times using the Transcreener® ADP Assay View Now