Search and Browse
Download
Enter Data
|
Assay Method Information |
|
| | Inhibition Kinetics Assay |
| Description: | Full length porcine calpain (156 nM), or papain (236 pM) was added to a solution of 100 mM NaCl, 50 mM HEPES, pH 7.6, 1 mM TCEP, 30 μM Suc-LLVY-AMC (SEQ ID NO: 8) substrate, and inhibitor (0.5 to 50 μM). Calpain reactions also contained CaCl2 (1 mM and 100 mM for porcine and rat respectively). Both substrate and inhibitors were dissolved in acetonitrile/DMSO (1:1) with the exception of E-64, dissolved in water. Organic solvent remained <2% in all reactions, and most often <1%. Reactions were carried out in microtiter 96-well plates, with 150 μL per well, 30° C., and product formation was monitored over time by fluorescence (Ex/Em 346/444 nm, with 420 nm cutoff filter). Kinetic values of kobs were determined via non-linear regression using one-phase association analysis and linear plots of 1/kobs vs. 1/[I] provided kinetic constants ki and KI. |
| Affinity data for this assay | |
|---|---|
| If you find an error in this entry please send us an E-mail | |
|
Home |
| |
Search |
| |
Deposit |
| |
SiteMap |
| |
About us |
| |
Email us |
| |
Info |
|
©2000 BindingDB. All rights reserved. |
|||||||||||||