Time-gated autofluorescence microscopy of motile green microalga in an optical trap

Abstract

Ultrafast time-gated fluorescence imaging of optically trapped single motile cells is presented. The biflagellar green microalga Haematococcus pluvialis was confined with picoNewton trapping forces in the focal volume of a high numerical aperture objective by near infrared multitraps. Trapping radiation of 100 mW power at the sample was provided by a Nd:YLF laser (1047 nm) operating in the cw mode. Simultaneously, cellular autofluorescence was excited with a 633 nm picosecond 80 MHz laser diode. An ultrafast gated intensified slow scan CCD camera system with a tunable gate width (200 ps-1 ms) and tunable time-delay (0-20 ns) between excitation and detection was used as fluorescence detector. We demonstrate fluorescence imaging of high temporal (sub-ns) and high spatial (sub-microm) resolution and fluorescence lifetime determination of intracellular autofluorescence based on chlorophyll excitation. Exposure to the herbicide DCMU resulted in an increase of fluorescence intensity and lifetime by 250% and 150%, respectively.