Multi-photon fluorescence microscopy - the response of plant cells to highintensity illumination

Multi-photon fluorescence microscopy has been cited for its advantage in in creased depth penetration due to low linear absorption and scattering coeff icient of biological specimen in the near infrared (NIR) range. Because of the need of high peak power for efficiently exciting two-photon fluorescenc e, the relationship between cell damage and peak power has become an intere sting and much debated topic in the applications of multi-photon fluorescen ce microscopy. It is conceivable that at high illumination intensity, non-l inear photochemical processes have impacts on cell physiology and viability in ways much different from low illumination in the linear domain. In this article, we discuss some of the issues in two-photon fluorescence microsco py, including the degree of transparency of the specimen, a comparison of s ingle- and two-photon excited fluorescence spectra, and the cell damage und er high intensity illumination, using plant cells as a model. (C) 2001 Else vier Science Ltd. All rights reserved.