Comparison of three‐dimensional imaging properties between two‐photon and single‐photon fluorescence microscopy

SUMMARY The imaging performance in single‐photon (1‐p) and two‐photon (2‐p) fluorescence microscopy is described. Both confocal and conventional systems are compared in terms of the three‐dimensional (3‐D) point spread function and the 3‐D optical transfer function. Images of fluorescent sharp edges and layers are modelled, giving resolution in transverse and axial directions. A comparison of the imaging properties is also given for a 4Pi confocal system. Confocal 2‐p 4Pi fluorescence microscopy gives the best axial resolution in the sense that its 3‐D optical transfer function has the strongest response along the axial direction.