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HTRF Kinase Binding Buffer, 20 ml

The Kinase Binding Buffer is a spare part intended for use in the HTRF® Kinase Binding platform only.

For research use only. Not for use in diagnostic procedures. All products to be used in accordance with applicable laws and regulations including without limitation, consumption and disposal requirements under European REACH regulations (EC 1907/2006).

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Feature Specification
Application Biochemical Enzymatic Assay

The Kinase Binding Buffer is a spare part intended for use in the HTRF® Kinase Binding platform only.

For research use only. Not for use in diagnostic procedures. All products to be used in accordance with applicable laws and regulations including without limitation, consumption and disposal requirements under European REACH regulations (EC 1907/2006).

Click to copy promo code to clipboard.
SAVE 15% NOW on online orders. Use promo code below.
YEAREND15
Offer valid until 12/15. Terms and conditions apply.
Product Variants
Unit Size: 20 ml
Part #:
62KBBRDD
List Price
USD 200.36
Unit Size: 200 ml
Part #:
62KBBRDF
List Price
USD 317.85

Overview

​The Kinase Binding Buffer is used to perform saturation binding (Kd-determination) and/or competitive binding assays within the HTRF® Kinase Binding platform only.

Specifications

Application
Biochemical Enzymatic Assay
Brand
HTRF
Detection Modality
HTRF
Product Group
Buffer
Shipping Conditions
Shipped Ambient
Target Class
Kinases
Technology
TR-FRET
Therapeutic Area
Metabolism/Diabetes
Neuroscience
Oncology & Inflammation
Unit Size
20 ml

Video gallery

How it works

Kd determination assay principal

The binding of a fluorescent tracer is detected in a sandwich assay format using either a specific Anti-GST, 6HIS antibody or Streptavidin labeled with Europium Cryptate (donor), which bind to the tagged Kinase, and a red fluorescent tracer labelled with d2 (acceptor). The detection principle is based on HTRF® technology. The HTRF ratio (665/620) will increase upon the addition of more tracer, and will saturate depending on the dissociation constant (Kd ) of the tracer to the tagged kinase. When an inhibitor of the kinase is added, the tracer will be displaced and the HTRF signal will disappear, depending on the dose.

kinase-binding-how-it-works-assay-principal-saturation-staurosporine-red-62kb01redc-62kb01rede.svg

 

IC50/Ki determination assay protocol

​Pharmacological evaluation of inhibitors of interest can be run in 96- or 384-well plates.

First, a dilution series of inhibitor ranging between 40 µM and 0.23 nM is prepared, and 5 µL of each concentration are dispensed into the plate. Next, 5 µL of tagged-Kinase are added, followed by 5 µL of anti-tag Eu-cryptate. Finally, 5 µL of tracer solution are added, prepared at 4x the final concentration. The HTRF ratio is measured after 1 H of incubation.

Analyses of the data give typical dose response curves ranging between 10 µM and 56 pM, enabling an evaluation of the IC50/Ki values for the inhibitor of interest.

kinase-binding-how-it-works-assay-protocol-inhibition-staurosporine-red-62kb01redc-62kb01rede.svg

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