Combined two-photon excitation and d→f energy transfer in a water-soluble Ir(III)/Eu(III) dyad: two luminescence components from one molecule for cellular imaging.

The first example of cell imaging using two independent emission components from a dinuclear d/f complex is reported. A water-stable, cell-permeable Ir(III) /Eu(III) dyad undergoes partial Ir→Eu energy transfer following two-photon excitation of the Ir unit at 780 nm. Excitation in the near-IR region generated simultaneously green Ir-based emission and red Eu-based emission from the same probe. The orders-of-magnitude difference in their timescales (Ir ca. µs; Eu ca. 0.5 ms) allowed them to be identified by time-gated detection. Phosphorescence lifetime imaging microscopy (PLIM) allowed the lifetime of the Ir-based emission to be measured in different parts of the cell. At the same time, the cells are simultaneously imaged by using the Eu-based emission component at longer timescales. This new approach to cellular imaging by using dual d/f emitters should therefore enable autofluorescence-free sensing of two different analytes, independently, simultaneously and in the same regions of a cell.