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HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
Generic Vial HTRF
| Feature | Specification |
|---|---|
| Application | Biochemical Enzymatic Assay |
Product variant
Unit Size: 1 vial
Part #:
61SEBALB
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).
HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
Generic Vial HTRF
HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
Product information
Overview
Some Tyrosine kinase enzymatic activity may require further assay optimization for a sensitive detection of phosphorylation levels. The Supplemented Enzymatic Buffer is a substrate stabilizing buffer that may be used to achieve an optimal enzymatic step prior to detection with the HTRF KinEASE TK kit, while maintaining inhibiting compound pharmacological characteristics.
Analytical performance
Supplemented Enzymatic Buffer performance
The HTRF KinEASE TK reaction may be run in the presence of SEB, a proprietary substrate stabilizing buffer. This buffer greatly enhances S/B and enables the use of lower kinase concentrations per well. Among validation on other tyrosine kinases, the case below shows the impact of SEB on tests conducted with Abl (Merck Millipore 14-459).
In this SEB titration run, Abl used at 5 ng/well, was incubated 30 min with substrate-biotin (1 µM), and a non-limiting ATP concentration (100 µM) in a final assay volume of 20 µl. SEB concentrations ranging from 0 nM to 50 nM were tested. As shown here, SEB enhances S/B quite significantly.
In this second experiment, Abl was used at concentrations ranging from 0.10 to 10 ng/ml under similar substrate and ATP conditions as previously described, and with either 2.5 or 5 nM SEB. The subsequent S/B improvement at increasing SEB levels makes the use of lower tyrosine kinase concentration possible.
| Tyrosine KinEASE assay | |||
|---|---|---|---|
| Enzyme | Compound | With SEB 50 nM | Without SEB |
| Abl | Staurosporine Gleevec |
0.108 0.043 |
- - |
| CSK | Staurosporine Nexavar |
0.493 4.220 |
0.417 6.175 |
| AGFR | Staurosporine Iressa |
1.069 0.001 |
0.861 0.002 |
| JAK2 | Staurosporine Sutent |
0.00030 0.800 |
0.00035 0.778 |
| JAK3 | Staurosporine Sutent |
0.001 7.639 |
0.001 4.597 |
| PTK5 | Staurosporine Nexavar |
0.039 > 0.1 |
0.036 > 0 .1 |
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
NASH/Fibrosis
Oncology & Inflammation
|
| Unit Size |
1 vial
|
Video gallery
HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
HTRF Supplemented Enzymatic Buffer, 12.5 nmoles
Resources
Are you looking for resources, click on the resource type to explore further.
Application Note
Got Kinase Targets? KinEASE is here to help
Measuring kinase activity under conditions you want
Download this complimentary Application Note to explore the features of the...
Guide
HTRF solutions, guide to major applications
This guide provides you an overview of HTRF applications in several therapeutic areas.
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