Most molecules absorb parts of white light, allowing us to see them with the naked eye. However, absorption contrast is not widely used for optical imaging of molecules because of the poor sensitivity involved.
On the other hand, fluorescence microscopy, which is based upon emission rather than absorption, is a much more sensitive technique. When molecules or atoms absorb energy they enter an excited state then spontaneously decay back to lower energy levels, in some cases giving rise to fluorescence. The problem is, not all chromophores fluoresce upon decay and instead they may take a non-radiative pathway. Such molecules must either be studied using the low sensitivity optical techniques or they need to be tagged using a fluorescent marker.
In a new technique, molecules of the sample are subjected to a pulse with the right energy and timing to enable another form of emission known as stimulated emission. In this case, emission competes with nonradiative decay such that it becomes the dominant energy pathway back to lower, more stable energy levels. Stimulated emission offers a sensitivity that is several orders of magnitude higher than normally seen for molecules that do not fluoresce. Team leader Xiaoliang Sunney Xie explains the technique's potential in the future of medicine. “It allows spectroscopic identification of molecules in living organisms and is free from the complication of light scattering by the sample. This opens many new possibilities for biomedical imaging, such as label free mapping of drug distributions and blood vessels in tissues.”
In fact, the team has already demonstrated proof-of-principle by imaging the delivery pathway of a drug that is normally difficult to study because its fluorescence is quenched on binding with tissues. The images obtained show the drug at different depths of the skin and can also confirm the hypothesis that the drug binds to cytoplasmic RNA to initiate cell death in cancerous cells.