Harder, Daniel; Fotiadis, Dimitrios (2012). Measuring substrate binding and affinity of purified membrane transport proteins using the scintillation proximity assay. Nature protocols, 7(9), pp. 1569-78. London: Nature Publishing Group 10.1038/nprot.2012.090
Full text not available from this repository.The scintillation proximity assay (SPA) is a rapid radioligand binding assay. Upon binding of radioactively labeled ligands (here L-[(3)H]arginine or D-[(3)H]glucose) to acceptor proteins immobilized on fluoromicrospheres (containing the scintillant), a light signal is stimulated and measured. The application of SPA to purified, detergent-solubilized membrane transport proteins allows substrate-binding properties to be assessed (e.g., substrate specificity and affinity), usually within 1 d. Notably, the SPA makes it possible to study specific transporters without interference from other cellular components, such as endogenous transporters. Reconstitution of the target transporter into proteoliposomes is not required. The SPA procedure allows high sample throughput and simple sample handling without the need for washing or separation steps: components are mixed in one well and the signal is measured directly after incubation. Therefore, the SPA is an excellent tool for high-throughput screening experiments, e.g., to search for substrates and inhibitors, and it has also recently become an attractive tool for drug discovery.
Item Type: |
Journal Article (Original Article) |
|---|---|
Division/Institute: |
04 Faculty of Medicine > Pre-clinic Human Medicine > Institute of Biochemistry and Molecular Medicine |
UniBE Contributor: |
Harder, Daniel, Fotiadis, Dimitrios José |
ISSN: |
1754-2189 |
Publisher: |
Nature Publishing Group |
Language: |
English |
Submitter: |
Factscience Import |
Date Deposited: |
04 Oct 2013 14:37 |
Last Modified: |
05 Dec 2022 14:11 |
Publisher DOI: |
10.1038/nprot.2012.090 |
PubMed ID: |
22864198 |
Web of Science ID: |
000308526300001 |
URI: |
https://boris.unibe.ch/id/eprint/15047 (FactScience: 222201) |
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