– CELT-327 is a potent, selective fluorescent hA2B/A3 adenosine receptor antagonist, exhibiting K_i values of 35.6 nM at A2B and 45.7 nM at A3 receptors. It efficiently labels HCT116 cells expressing A2B adenosine receptors in a dose-dependent manner, demonstrating rapid, uniform spread and integration of fluorescence within cell monolayers. In live HCT116 spheroids, it achieves consistent staining throughout, penetrating even the deepest layers. As a fluorescent adenosine receptor probe, CELT-327 is ideal for applications in fluorescence binding assays, live cell imaging, flow cytometry, and fluorescence polarization assays, serving as an effective alternative to GPCR radiolabeling ligands. Its excitation and emission peaks are at 589 nm and 616 nm, respectively, making it suitable for TR-FRET assays as an acceptor dye, particularly in combination with the CoraFlor1 TR-FRET donor.