Optimization of laser beams in FRAP experiments of microscopical objects

In fluorescence recovery after photobleaching (FRAP) experiments the sample is irradiated on a small spot, the diameter of which must be related to the sample size and the diffusion constant to be measured. This paper considers the conventional FRAP set-up where a laser beam is directed through a microscope vertical illuminator to the sample. The requirements of an intermediate optical system producing a Gaussian beam with a waist of given radius in the microscope object plane are considered, and the optical parameters determined.