Oxygen-Insensitive Aggregates of Pt(II) Complexes as Phosphorescent Labels of Proteins with Luminescence Lifetime-Based Readouts

The synthesis and photophysical properties of a tailored Pt(II) complex are presented. The phosphorescence of its monomeric species in homogeneous solutions is quenched by interaction with the solvent and therefore absent even upon deoxygenation. However, aggregation-induced shielding from the environment and suppression of rotovibrational degrees of freedom trigger a phosphorescence turn-on that is not suppressed by molecular oxygen, despite possessing an excited-state lifetime ranging in the microsecond scale. Thus, the photoinduced production of reactive oxygen species is avoided by the suppression of diffusion-controlled Dexter-type energy transfer to triplet molecular oxygen. These aggregates emit with the characteristic green luminescence profile of monomeric complexes, indicating that Pt-Pt or excimeric interactions are negligible. Herein, we show that these aggregates can be used to label a model biomolecule (bovine serum albumin) with a microsecond-range luminescence. The protein stabilizes the aggregates, acting as a carrier in aqueous environments. Despite spectral overlaps, the green phosphorescence can be separated by time-gated detection from the dominant autofluorescence of the protein arising from a covalently bound green fluorophore that emits in the nanosecond range. Interestingly, the aggregates also acted as energy donors able to sensitize the emission of a fraction of the fluorophores bound to the protein. This resulted in a microsecond-range luminescence of the fluorescent acceptors and a shortening of the excited-state lifetime of the phosphorescent aggregates. The process that can be traced by a 1000-fold increase in the acceptor's lifetime mirrors the donor's triplet character. The implications for phosphorescence lifetime imaging are discussed.