Current activities on our research of soft x-ray lasers and higher harmonic generations using intense, subpicosecond lasers are reviewed. Especially for soft x-ray lasers we experimentally demonstrate that by longitudinally pumping 2-mm-long molybdenum preformed plasma with high-intensity 475 fs duration laser pulse, a highly directive soft x-ray laser at 18.9 nm wavelength is generated. The divergence of the beam is evaluated to be of the submillirad order, and only requires pump laser energy of 150 mJ. Simulations show that the pedestal in the main pump pulse can generate electron density and gain profiles with large spatial gradients, which result in the selective amplification of low-order transverse modes. The present result is the demonstration of an efficient and alternative method of improving the spatial coherence of x-ray lasers with amplified spontaneous emission medium, with possibilities of becoming an excellent tool to explore various application experiments. In higher harmonic generation, topics related to observation of blue shift due to collisionless absorption process is described. We demonstrate the generation of high harmonics (up to the 65^th order, λ=12.24 nm) of a Ti:sapphire laser radiation after the propagation of femtosecond laser pulses through the low-excited boron plasma produced by a prepulse radiation on the surface of different targets. High-order harmonics generated from the surface plasma of most targets showed a plateau pattern. The harmonic generation in these conditions assumed to occur due to the interaction of femtosecond pulses with ions. The conversion efficiencies in the plateau region were varied between 10^-7 to 8×10^-5 depending on the target. The main contribution to the limitation of harmonic generation efficiency and cutoff energy was attributed to the free-electrons-induced self-defocusing of main pulse.

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We measured the decay time shortening of the FRET(Fluorescence Resonance Energy Transfer) signals in a mixed donor-acceptor protein pair such as the cameleon calcium ion indicator and a ligand activated GRIN-Go proteins pair with a newly improved an ultra fast time-resolved FRET spectroscopy system using photon counting method in this study. G protein-coupled receptors is the most important protein family for the recognition of many chemical substances at the cell surface. For both protein pair systems, donor CFP protein fluorescence lifetime curves showed the double decay and the fluorescence lifetime distinctly shortened in the FRET condition. Simultaneously we got the emission spectral distribution of luminous protein within 10ns from excitation, which is the first data acquired time-resolvedly at the initial stage in the FRET condition. This system enables us to trace the dynamics of the interaction between proteins at the ligand induced activated state, molecular structure change and combination or dissociation.

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Recent progress of the optical microscopy, especially the nearfield microscopy, has enabled us to study functions and interactions of protein molecules at single molecular level. Single molecule imaging has become an indispensable technique to life science research. On one hand, after the determination of the whole DNA sequences of human genome, the remained issues are elucidation of the functions of genes. Single-molecule imaging technique is expected to greatly contribute to such problems. Here, I review and survey the functional analysis of biomolecules by single-molecule imaging and manipulation.

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In this article, we describe a noninvasive, straightforward methods to inactivate selected proteins in living cells with high spatiotemporal resolution (MP-CALI). In MP-CALI, a target protein is tagged with enhanced green fluorescent protein (EGFP). When EGFP is excited via two photon excitation process by a femtosecond pulse laser, reactive oxygen species (ROS) are generated and the protein of interest is specifically inactivated by ROS. MP-CALI should find broad applications and provide valuable information on protein functions.

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Fluorescent imaging is a useful tool for monitoring spatiotemporal dynamics of intracellular signals and understanding of a lot of biological phenomena. Recently, multicolor imaging technique has been highlighted, using many color variation of fluorescent molecules. This commentary article has a brief summary for development of colorful variants of fluorescent protein and quantum dots sustaining multicolor imaging. We also review optical systems for multicolor imaging, discussing the flexibility of excitation and detection lights and the temporal and spatial consistency of detected images. Though overlapping emission spectra limits the number of fluorescent molecules that can be detected simultaneously, some computational methods have been proposed to overcome this issue. We introduce our preliminary work for live cell multicolor imaging and review the numerical solutions for multicolor imaging.

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