Fluctuation of probe beam in thermolens schematics as potential indicator of cell metabolism, apoptosis, necrosis and laser impact

摘要

Recently we discovered that in developed integrated flow cytometry using combination high resolution transmission and two beam (pump-probe) of photothermal microscopy with flow module the temporal fluctuation of probe beam even without pump laser pulse are very sensitive to functional states of single cells (e.g. with 2-3 differences in average fluctuation amplitude for live and dead cells). In this paper we are focusing on theoretical and experimental studies of these new phenomena using thermolens schematics with highly stabilized continuous-wave He-Ne laser and photodiode/photomultiplier with pinhole. This scheme allows to study both random and laser-induced thermolens effects in passive and active mode including short and long term monitoring of scattered light fluctuation in trans-illumination mode at single cellular and even sub-cellular levels in vitro, and in vivo. We demonstrate these schematics has some potential to monitor intracellular dynamics including change in cell metabolism, and necrosis, especially, under pump laser impacts in vitro in stationary condition. Besides, we present some relevant data obtained with other similar schematics, including evaluation of bacterial motility by dynamic light scattering. We discuss further potential developing this approach for flow cytometry with fast speed photodetector and digital camera and the multi- wavelength statistical and correlation analysis of speckle-related signals in applications to analysis of complex motions and functional properties of cells in bioflows including the evaluation of the rotational and translational components of the motion of individual live and dead cells.