Development of a fluorescent ligand-binding assay using the AcroWell filter plate.
One of the most powerful tools for receptor research and drug discovery is the use of receptor-ligand affinity screening of combinatorial libraries. Early work involved the use of radioactive ligands to identify a binding event; however, there are numerous limitations involved in the use of radioactivity for high throughput screening. These limitations have led to the creation of highly sensitive, nonradioactive alternatives to investigate receptor-ligand interactions. Pall Gelman Laboratory has introduced the AcroWell, a patented low-fluorescent-background membrane and sealing process together with a filter plate design that is compatible with robotic systems. Taken together, these allow the AcroWell 96-well filter plate to detect trace quantities of lanthanide-labeled ligands for cell-, bead-, or membrane-based assays using time-resolved fluorescence. Using europium-labeled galanin, we have demonstrated that saturation binding experiments can be performed with low-background fluorescence and signal-to-noise ratios that rival traditional radioisotopic techniques while maintaining biological integrity of the receptor-ligand interaction. In addition, the ability to discriminate between active and inactive compounds in a mock galanin screen is demonstrated with low well-to-well variability, allowing reliable determination of positive hits even for low-affinity interactions.
Duke Scholars
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Related Subject Headings
- Receptors, Neuropeptide
- Receptors, Galanin
- Radioligand Assay
- Medicinal & Biomolecular Chemistry
- Ligands
- Iodine Radioisotopes
- Humans
- Galanin
- Fluorescent Dyes
- Fluorescence
Citation
Published In
DOI
ISSN
Publication Date
Volume
Issue
Start / End Page
Location
Related Subject Headings
- Receptors, Neuropeptide
- Receptors, Galanin
- Radioligand Assay
- Medicinal & Biomolecular Chemistry
- Ligands
- Iodine Radioisotopes
- Humans
- Galanin
- Fluorescent Dyes
- Fluorescence