In this communication, we show that off-axis digital holography combined to phase-shifting acquisition of holograms
is an effective microscopic tool to fully localize, in three dimensions, transmembrane receptors of living
cells tagged with Gold nanocolloids. Gold nanoparticles, known for their interesting optical properties as well as
for their noncytotoxicity are used here as biomarkers to target the cellular receptors.
We have developed a holographic optical tweezers system based on diffractive optical elements (DOES) implemented
on a liquid crystal spatial light modulator (LC-SLM) able to generate fine positioned traps on the sample. Our own
algorithms and code allows to calculate phase DOES that can transform a single laser beam into an array of independent
traps, each with individually specified characteristics, arranged in arbitrary three-dimensional (3D) geometrical
configurations. Different DOEs can be dynamically projected to the SLM in order to achieve a rearrangement of the
configuration of the trapping spots. Silica or latex micro-beads are trapped in different configurations of spots to
demonstrate the fine control capability on each trap. Our setup is built on a standard video microscope coupled with a
laser source, a spatial light modulator and a three axis nano-positioning system. It allows to obtain 3D multi-trapping
and a fine calibration for the positioning of the traps.
Novel MCP detectors for time- and space-correlated single photon counting (TSCSPC) spectroscopy, featuring delay-line (DL) or quadrant anode (QA), are employed in microscopic fluorescence lifetime imaging on the picosecond time scale. The linear DL-MCP-PMT is characterized by a spatial instrument response function (IRF) of 100 micrometer FWHM, resulting in 200 space channels, whereas the QA-MCP-PMT is a 2D imager with 400 by 400 pixel at 40 micrometer resolution. The detectors have a temporal IRF of 75 ps (DL) and 120 ps (QA) FWHM, sufficient for 10 ps time resolution. First results on TOTO-fixed cell systems are presented, demonstrating high-quality kinetics at subcellular resolution, with up to 6 lifetime species at higher dye concentrations, characteristically distributed among individual cell compartments. A comparison with TOTO/DNA- suspensions is made that serve as reference system.