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Development of a quantitative fluorescence-based ligand-binding assay.

ABSTRACT: A major goal of biology is to develop a quantitative ligand-binding assay that does not involve the use of radioactivity. Existing fluorescence-based assays have a serious drawback due to fluorescence quenching that accompanies the binding of fluorescently-labeled ligands to their receptors. This limitation of existing fluorescence-based assays prevents the number of cellular receptors under investigation from being accurately measured. We have developed a method where FITC-labeled proteins bound to a cell surface are proteolyzed extensively to eliminate fluorescence quenching and then the fluorescence of the resulting sample is compared to that of a known concentration of the proteolyzed FITC-protein employed. This step enables the number of cellular receptors to be measured quantitatively. We expect that this method will provide researchers with a viable alternative to the use of radioactivity in ligand binding assays.

SUBMITTER: Breen CJ

PROVIDER: S-EPMC4861924 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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Development of a quantitative fluorescence-based ligand-binding assay.

Breen Conor J CJ Raverdeau Mathilde M Voorheis H Paul HP

Scientific reports 20160510

A major goal of biology is to develop a quantitative ligand-binding assay that does not involve the use of radioactivity. Existing fluorescence-based assays have a serious drawback due to fluorescence quenching that accompanies the binding of fluorescently-labeled ligands to their receptors. This limitation of existing fluorescence-based assays prevents the number of cellular receptors under investigation from being accurately measured. We have developed a method where FITC-labeled proteins boun ...[more]

PMID: 27161290

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Dataset Information

Development of a quantitative fluorescence-based ligand-binding assay.

Publications

Development of a quantitative fluorescence-based ligand-binding assay.

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