Assay development and data analysis of receptor-ligand binding based on scintillation proximity assay.
Metab Eng
; 7(1): 38-44, 2005 Jan.
Article
en En
| MEDLINE
| ID: mdl-15721809
ABSTRACT
In this paper, we described the optimization of a generic binding assay to measure ligand-receptor interactions for peroxisome proliferator-activated receptors (PPARs). The assay is based on scintillation proximity assay, in which a protein is coated on scintillant-incorporated beads, and a radiolabeled ligand stimulates the beads to emit a signal by binding to the immobilized protein. An intrinsic binding affinity of unlabeled ligands is determined by competitive displacement of the radioligand. The protein coating and ligand binding are achieved in one step by simply mixing ligands, protein and beads in sequence. No additional steps of pre-coating and washing of beads are required. Protein is captured on beads effectively by electrostatic interactions, thus no affinity labeling of protein is required. In data analysis, ligands are grouped into two classes based on their binding affinities. For tight binding ligands, an equation is derived to accurately determine the binding affinity. Otherwise a general equation applies. This quantitative and high throughput assay provides a tool to screen a large library of molecules in search of potent ligands.
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Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
Conteo por Cintilación
/
Radioinmunoensayo
/
Mapeo de Interacción de Proteínas
/
PPAR gamma
Tipo de estudio:
Diagnostic_studies
/
Evaluation_studies
Idioma:
En
Revista:
Metab Eng
Asunto de la revista:
ENGENHARIA BIOMEDICA
/
METABOLISMO
Año:
2005
Tipo del documento:
Article
País de afiliación:
Estados Unidos