| Autor: |
Maris-Johanna Tahk, Tõnis Laasfeld, Elo Meriste, Jose Brea, Maria Isabel Loza, Maria Majellaro, Marialessandra Contino, Eddy Sotelo, Ago Rinken |
| Jazyk: |
angličtina |
| Rok vydání: |
2023 |
| Předmět: |
|
| Zdroj: |
Frontiers in Molecular Biosciences, Vol 10 (2023) |
| Druh dokumentu: |
article |
| ISSN: |
2296-889X |
| DOI: |
10.3389/fmolb.2023.1119157 |
| Popis: |
Dopamine receptors are G-protein-coupled receptors that are connected to severe neurological disorders. The development of new ligands targeting these receptors enables gaining a deeper insight into the receptor functioning, including binding mechanisms, kinetics and oligomerization. Novel fluorescent probes allow the development of more efficient, cheaper, reliable and scalable high-throughput screening systems, which speeds up the drug development process. In this study, we used a novel Cy3B labelled commercially available fluorescent ligand CELT-419 for developing dopamine D3 receptor-ligand binding assays with fluorescence polarization and quantitative live cell epifluorescence microscopy. The fluorescence anisotropy assay using 384-well plates achieved Z’ value of 0.71, which is suitable for high-throughput screening of ligand binding. The assay can also be used to determine the kinetics of both the fluorescent ligand as well as some reference unlabeled ligands. Furthermore, CELT-419 was also used with live HEK293-D3R cells in epifluorescence microscopy imaging for deep-learning-based ligand binding quantification. This makes CELT-419 quite a universal fluorescence probe which has the potential to be also used in more advanced microscopy techniques resulting in more comparable studies. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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