Microscopy System for Distinguishing Stimulated Emissions as a Means of Increasing Signal

The invention pertains to a system and method for distinguishing stimulated emissions as a means of enhancing signal strength of fluorescent markers in fluorescence microscopy applications. The system is arranged such that an excitation beam (e.g., laser beam) illuminates a sample along some axis exciting the fluorescent markers used in the sample. A second light beam, a stimulation beam, illuminates the sample along another axis, possibly the same as that of the excitation beam. It has been found that if the excited fluorescent molecules are illuminated with light of a stimulation beam at a particular wavelength after initial excitation, the fluorescent molecules will emit light at this wavelength that can be separately detected. An excited fluorescent molecule may be stimulated by light at a wavelength different from the initial excitation beam to boost the signal. The stimulated emission then generated by the fluorescent molecules travels along the same access as the stimulation beam and, as such, the system is configured by a stimulation beam block component associated with an objective lens that prevents or reduces stimulation beam detection but allows detection of the stimulated emission. Another way the invention achieves this is by refocusing both the excitation and stimulation beams through capture by an excitation objective. A filter is then used to filter out light focused by the excitation objective from the simulated emission sent back by the fluorescent molecule.