Measure Enzymatic Activity with the Transcreener® ADP² Assay Kit

The Transcreener ADP2 Assay uses a highly selective antibody and far-red fluorescent tracer to directly detect ADP produced by any ADP-producing enzyme (i.e. kinases, ATPases, helicases, carboxylases, nucleotidases, and chaperonins). The direct detection method offers a unique approach that simplifies the protocol (no coupling enzyme needed) and reduces compound interference. The assay can be used to:

  • Measure activity of any ADP-producing enzyme
  • Screen compound libraries for modulators
  • Quantify inhibitor potency
  • Inhibitor selectivity profiling
  • Measure drug-target residence time

Overview Transcreener ADP Assay

Image Credit: BellBrook Labs

Targets Validated with the Transcreener ADP2 Assay

BellBrook Labs develops complete assay solutions for tougher enzyme targets that are difficult to source enzymes and/or develop assays for (so you can focus on optimizing your lead molecules). Currently, they have assay solutions for the ADP-producing enzyme targets listed in the table below. Click on a target to learn more. 

Enzyme
ADK RIPK1
AMPK RIPK2
IKK-beta TBK1
IRAK4 p97
JAK1 Ketohexokinase
JAK3 NSP13
MAPK1 POLQ
MAPK8 RIG-I
PKR MDA5
SRC-Kinase WRN Helicase

Source: BellBrook Labs

Features

  • Direct detection of unlabeled ADP
  • Easy to use and HTS-ready
  • Highly sensitive assay reduces enzyme usage
  • Generic nature makes it compatible with any ADP-producing enzyme
  • Robust Assay Z' > 0.7 under initial velocity conditions
  • Far-red fluorescent readouts minimize compound interference
  • A safe, non-radioactive method
  • Use with ATP concentrations from 0.1 to 1000 µM
  • Available in FP, FI, or TR-FRET readouts 
  • Can be used in continuous or endpoint mode

Example Data using MAPK1 Kinase

Robust Assay Ready to Scale for HTS

Z

Z' = 0.90. Robust data is neccessary to confirm the assay is well-suited for screening large compound libraries in a high-throughput format. 

Image Credit: BellBrook Labs

Detection of ADP Under Initial Velocity Conditions

MAPK1 Enzyme Titration with Transcreener ADP FP Assay

MAPK1 enzyme titration using Transcreener ADP2 Assay with FP readout. The enzyme buffer included 20 mM Tris pH 7.5, 10 mM MgCl2, 1 mM DTT, and 0.01% Tween-20. The detection mix included 1X Stop and Detect Buffer B, 6.4 μg/mL ADP2 Antibody, and 4 nM ADP2 Tracer. 

Image Credit: BellBrook Labs

MAPK1 Linear Response with Transcreener ADP FP Assay

MAPK1 linear response. The assay shows linearity when raw data is converted to ADP using a standard curve. 

Image Credit: BellBrook Labs

Screen Compound Libraries & Perform Dose Responses

MAPK1 Dose Response with Transcreener ADP FP Assay

MAPK1 dose response with compound staurosporine using Transcreener ADPFP Assay. IC50 (Staurosporine)= 0.77 μM. 

Image Credit: BellBrook Labs

Lead Discovery Services

BellBrook Labs also offers lead discovery services for scientists who want to advance their program but don't want to bring a biochemical assay in-house. Their services feature customizable assay conditions, quick turnaround times, and direct collaboration with a BellBrook scientist. Lead discovery services can include:

  • Inhibitor Screening
  • Inhibitor Potency Profiling
  • Inhibitor Selectivity Profiling
  • Residence Time Measurements
  • Mechanism of Action Studies
  • Triage Non-Stoichiometric Inhibitors
  • Compound-Target Binding with Thermal Shift Assays 

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