Efficient simulation of autofluorescence effects in microscope lenses.

The use of fluorescence in microscopy is a well-known technology. Due to autofluorescence in the materials of optical components, the contrast of the image is degraded. The calculation of autofluorescence is usually performed by brute-force methods such as the Monte Carlo-based volume scattering. The efficiency of calculations in this case is extremely low, and a huge number of rays must be calculated. In stray light calculations, the concept of important sampling is used to reduce computational effort. The idea is to calculate only rays, which have the chance to reach the target surface. The fluorescence conversion can be considered to be a scatter process, and therefore a modification of this idea is used here. The reduction factor is calculated by comparing the size of the illuminated phase space domain with the corresponding acceptance domain in every z plane of the lenses. The boundaries of the domains are determined by tracing the limiting rays of the light cone of the source as well as the pixel area under consideration. The small overlap of both domains can be estimated by geometrical considerations. The correct photometric scaling and the discretization of the volumes must be performed. The errors resulting from necessary approximations can be corrected without greatly increasing computational effort. The run time is reduced by a factor of 10. It is shown with some practical examples of microscope lenses that the results are comparable with conventional methods. Additionally, a quasi-analytical model that describes the dependence of autofluorescence on various lens parameters is derived.