A molecular clip combining a doubly substituted fluorescent anion–π donor probe and two flexible arms bearing H-bond motifs constitutes a new generation of anion receptors. Five simultaneous non-covalent interactions are highlighted by theoretical complexation studies with five different anions. A large range of analytical techniques (electrospray-tandem mass spectrometry, NMR, UV-visible, steady-state and time-resolved fluorescence) were deployed to evaluate the stoichiometry and association constants with the selected anions. The photophysical and anion–π donor properties of the tetrazine ring allowed fine characterization of the binding properties of the ligand. Based on previously published results, an anti-cooperativity effect in non-covalent interactions was demonstrated.