Non-Specific Binding of the Fluorescent B-Adrenergic Receptor Probe Alprenolol-NBD

Abstract

The fluorescent ß-adrenergic receptor probe alprenolol-NBD was found to exhibit a high affinity (K_d 3.2 nM) and a low capacity (10 fmol/mg protein) for the ß₂-adrenergic receptor on living Chang liver cells but also a high affinity (K_d 320 nM) for non-ß-adrenergic receptor binding sites with a very high capacity (28,000 fmol/mg protein). Calculations are presented which make clear that less than 3% of the binding of alprenolol-NBD during visualization experiments is ß-adrenergic receptor related. Furthermore, it is shown that besides the downregulation of ß-adrenergic receptors during incubation with isoproterenol, the high-affinity non-ß-receptor binding sites are also deminishing during incubation with isoproterenol. Based on our findings it is concluded that the results of Henis et al. (1) who claimed the visualization of the ß-adrenergic receptor population on Chang liver cells by alprenolol-NBD must be interpreted as an almost completely non-specific fluorescence.