Fluorescence resonance energy transfer (FRET) microscopy: a tool for in situ study of cellular structures

A dye pair characterized by favorable spectral properties allows a simplified analytical procedure, based on the measurements of both donor and acceptor emission in double-stained cytological samples, to be applied to evaluate both the relative efficiency of the energy transfer (FRET) process and its topological distribution. Propidium Iodide, a DNA intercalating agent, has been used in combination with Hoechst 33258, a non-intercalating dye specific for A-T sequences of DNA, to assess the chromatin arrangement in human fibroblasts in both quiescent (G0) and cycling (G1) phases. The results indicate that the cells in the two phases, that cannot be distinguished on the basis of the DNA content, exhibit differences of the FRET efficiency relative value that can be ascribed to chromatin structure modification related to gene activation processes.