new BindingDB logo
myBDB logout

Assay Method Information

Assay Name:  Functional Assay
Description:  Functional [35S]GTPgammaS binding assays were conducted as follows. kippa opioid receptor membrane solution was prepared by sequentially adding final concentrations of 0.026 ug/ul kippa membrane protein (in-house), 10 ug/mL saponin, 3 uM GDP and 0.20 nM [35S]GTPgammaS to binding buffer (100 mM NaCl, 10 mM MgCl2, 20 mM HEPES, pH 7.4) on ice. The prepared membrane solution (190 ul/well) was transferred to 96-shallow well polypropylene plates containing 10 ul of 20x concentrated stock solutions of agonist prepared in DMSO. Plates were incubated for 30 min at a temperature of about 25° C. with shaking. Reactions were terminated by rapid filtration onto 96-well Unifilter GF/B filter plates (Perkin Elmer, Shelton, Conn.) using a 96-well tissue harvester (Packard) and followed by three filtration washes with 2001 ice-cold binding buffer (10 mM NaH2PO4, 10 mM Na2HPO4, pH 7.4). Filter plates were subsequently dried at 50° C. for 2-3 hours.
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

 
Last update November 1, 2007
©2000 BindingDB. All rights reserved.