FFT-Based Method for Attenuation Correction in Fluorescence Confocal Microscopy

摘要

A problem in three dimensional imaging by a confocal scanning laser microscope inthe (epi)fluorescence mode is the darkening of the deeper layers due to absorption and scattering of both the excitation and the fluorescence light. A new method to correct for these effects is proposed. The approach, valid for weak attenuation, consists of multiplying the measured fluorescence intensity by a correction factor involving a convolution integral of the measured signal, which can be computed efficiently for the Fast Fourier Transform (FFT). Analytical and numerical estimates as to the degree of attenuation under which the method is valid are given and the method is tested on various test images. The method is compared with a recent iterative method with regard to numerical accuracy and computational efficiency.