I've been reading about pH and ion sensors that rely on a ratiometric approach, which supposedly circumvents many problems of intensity-based methods such as dye bleaching, dye leaching, and fluctuations in source intensity. I can think of situations though when this might not be the case.
For example, fura-2 is excited at two independent wavelengths, and the two emissions are detected sequentially. If the excitation source intensity at either wavelength were to fluctuate, the ratio of the emissions would be off from the calibration. Another example would be a FRET sensor in which the acceptor dye leaches at a faster rate than the donor dye; the emission ratio of acceptor to donor would no longer match the calibration.
So when does ratiometric measurement account for signal variations like dye bleaching, dye leaching, and fluctuations in source intensity? Is it only when there is one dye and one excitation source? Am I missing any other requisites? It seems like if there's only a single dye that bleaches/leaches or a single excitation source that fluctuates, the change in dye fluorescence would remain proportional such that the emission ratio remains the same.