Spectral imaging coupled to linear unmixing is becoming an important staple in the microscopist's toolbox, particularly when applied to the elimination of autofluorescence and for FRET investigations. Instruments equipped for spectral imaging are becoming increasingly popular and many confocal microscopes now offer this capability. Widefield fluorescence and brightfield microscopy are also being used more frequently for resolving complex fluorophore and absorbing dye mixtures, a trend that should continue into the future.