Our potent hA₁/A_2B adenosine receptor fluorescent antagonist shows affinity for hA₁ and hA_2B adenosine receptors (K_i = 1.89 nM and 24.65 nM respectively in radioligand binding assays), intermediate affinity for hA_2A (K_i = 80.33 nM) and very low affinity for hA₃ adenosine receptor (K_i = 967.48 nM). It allows to perform cell visualization in fluorescence microscopy, confocal microscopy and high content system experiments. It is potentially suitable for other fluorescence-based assays.