Analysis of cellular response to photodynamic treatment using digital holographic microscopy, SLIM, and FLIM

The response of cells of different origin to photodynamic treatment in vitro was studied using two realizations of quantitative phase imaging (QPI): off-axis digital holographic microscopy and spatial light interference microscopy (SLIM), and using fluorescence lifetime imaging microscopy (FLIM). Holographic techniques were shown to allow noninvasive monitoring and analysis of the response of both individual cells in a sample and their entire population to photodynamic treatment. Dynamics of changes in the phase shift introduced by cells provided information on cell death type and rate. Utilization of a low-coherence radiation source in the SLIM realization ensured reduced measurement error due to an absence of coherent noise. Changes in the fluorescence intensity and decay time of the applied chlorin-based photosensitizer in cells were shown to be due to photobleaching of the photosensitizer, rather than to intracellular processes occurring in the course of cell death. The observed variations in optical and morphological parameters of cells as a function of treatment dose were shown to conform to the specific cell death pathways. The advantages and disadvantages of each technique are discussed.