We demonstrate that incorporating Förster resonant energy transferring (FRET) mechanism to the Whispering gallery mode (WGM) microcavity can greatly enhance the sensitivity. A donor-dye doped microsphere is embedded in an acceptor dye solution, which forms a FRET-WGM sensing platform. Pumping the WGM microcavity with a pulsed laser, we obtained simultaneous lasing of donor and acceptor dyes. The gap between donor and acceptor resonant wavelengths serves as a readout for acceptor quantification. Compared with the pure WGMs sensing system, the detection limit of FRET-WGM is greatly decreased, which enables us to realize real-time monitoring and quantitative analysis of intracellular fluorescent substance at single cell level.
We briefly review the key technology of modern fiber based femtosecond laser sources summarizing advantages and
disadvantages of different mode-locking solutions. A description of possible extensions of a FemtoFiber-type modelocked
Er-doped fiber laser oscillator (1560 nm) reveals the flexibility with respect to wavelength coverage (488 nm ..
2200 nm) and pulse duration (10 fs .. 10 ps). The resulting FemtoFiber family and its versions for instrument integration
allow one to use these state-of-the-art light sources in many important applications, e.g. THz spectroscopy and
microscopy. We show that, depending on the fiber laser model and the THz emitter, THz radiation can be produced with
4-10 THz bandwidth and detected with up to 60 dB signal-to-noise ratio (SNR). Electronically controlled optical
scanning (ECOPS) - a unique method for fast, precise and comfortable sampling of the THz pulse or other pump-probe
experiments - is described and recommended for efficient data acquisition. As examples for modern microscopy with
ultrafast fiber lasers we present results of two-photon fluorescence, coherent microscopy techniques (SHG/THG/CARS)
and fluorescence lifetime imaging (FLIM).
Intrinsic Second Harmonic Generation (SHG) signals obtained from the motor protein myosin are of particular interest for 3D-imaging of living muscle cells. In addition, the new and powerful tool of 4Pi microscopy allows to markedly enhance the optical resolution of microscopy as well as the sensitivity for small objects because of the high peak intensities due to the interference pattern created in the focus. In the present study, we report, to our knowledge for the first time, measurements of intrinsic SHG signals under 4Pi conditions of type A. These measurements on mammalian myofibrilar structures are combined with very high resolution 4Pi fluorescence data obtained from the same preparations. We have chosen myofibrillar preparations of isolated mammalian muscle fibers as they (i) possess a regular repetitive pattern of actin and myosin filaments within sarcomers 2 to 3 μm in length, (ii) consist of single myofibrils of small total diameter of approximately 1 μm and (iii) are ideally suited to study the biomedically important process of force generation via calcium regulated motor protein interactions. Myofibrillar preparations were obtained from murine skeletal and heart muscle by using a combined chemical and mechanical fractionation1 (Both et al. 2004, JBO 9(5):882-892). BODIPY FL phallacidin has been used to fluorescently label the actin filaments.
The experiments were carried out with a Leica SP2 multi photon microscope modified for 4Pi measurements using a Ti:Sa laser tuned to 850-900 nm. SHG as well as fluorescence photons were detected confocally by a counting APD detector. The approach taken our study provides new 3D-data for the analysis and simulation of the important process of excitation-contraction coupling under normal physiological as well as under pathophysiological conditions.
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