Selection of viable cells with known DNA content

Cells doubly stained with Hoechst 33342 (H-33342) for DNA content and fluorescein diacetate for viability can be selected on the basis of both criteria using a single UV laser flow sorter. The selection is made possible due to resonance energy transfer occurring between the H-33342 and fluorescein fluorophores. Both a static fluorescence microscope and a dual laser flow sorter were used to demonstrate that energy transfer occurs in the doubly stained cells and that the sensitized emission in conjunction with the DNA emission can be used to select populations of cells with known DNA content and viability. The results indicate that fluorescein liberated by cellular esterases is freely accessible to the nucleus.