Radioligand Binding Assay
Radioligand binding assay can be used to characterize ligand binding to a receptor. This method allows to determine the affinity of a radioligand (KD) or a non-radioactive compound (IC50, Ki) as well as receptor density (Bmax) or mode of interaction (whether the compound is binding to orthosteric or allosteric site). Radioactivity can also be used for functional assays in order to determine the biological activity of a ligand (whether it is an agonist or antagonist), potencies (EC50) and efficacies (Emax). Additionally, kinetic measurements can be done do determine the rate constants of association and dissociation (kon and koff), but for these experiments we usually prefer fluorescence based methods, such as fluorescence anisotropy, which allows to measure ligand binding in real-time.
Currently we have a laboratory equipped for the work with radioactive isotopes 3H and 35S. We have the capacity to work with 20 ml tubes suitable for liquid scintillator counter Rack Beta (LKB). We also have a 24- and 96-well FilterMate Harvester (Perkin Elmer) for the filtration step and MeltiLex Heatsealer (Perkin Elmer) for impregnating solid scintillant. Together these allow us to work with microtiter plates suitable for luminescence counter Wallac MicroBeta TriLux (Perkin Elmer). This equipment has been successfully applied to study radioligand binding in different receptor preparations, starting from budded baculovirus particles (Allikalt and Rinken 2017) and cell membrane preparations (Reinart-Okugbeni et al. 2012, Reinart-Okugbeni et al. 2013, Dzirkale et al. 2013) to native tissues (Tekko et al. 2017).
Currently we have a laboratory equipped for the work with radioactive isotopes 3H and 35S. We have the capacity to work with 20 ml tubes suitable for liquid scintillator counter Rack Beta (LKB). We also have a 24- and 96-well FilterMate Harvester (Perkin Elmer) for the filtration step and MeltiLex Heatsealer (Perkin Elmer) for impregnating solid scintillant. Together these allow us to work with microtiter plates suitable for luminescence counter Wallac MicroBeta TriLux (Perkin Elmer). This equipment has been successfully applied to study radioligand binding in different receptor preparations, starting from budded baculovirus particles (Allikalt and Rinken 2017) and cell membrane preparations (Reinart-Okugbeni et al. 2012, Reinart-Okugbeni et al. 2013, Dzirkale et al. 2013) to native tissues (Tekko et al. 2017).