Laser scanning confocal microscopy cannot distinguish fluorescence signals of different labels, and there are overlapping and interference of different fluorescence signals in the acquired images. In order to solve this problem, a laser scanning confocal microscopy spectral imaging system based on filters was designed, to realize spectral screening and detection of different fluorescence signals. Spectral screening of the optical signals from the laser scanning confocal microscopy was performed using dichroic filters and detected through the blue-violet channel, the green channel, the yellow channel and the red-orange channel, respectively. The spectral imaging system operates in the wavelength range of 400nm≤λ≤720nm. The green channel and the yellow channel jointly achieve signal detection in the 475nm≤λ≤625nm wavelength range. In addition, the green channel and the yellow channel can further screen the signal spectrum through the continuously variable filters. The minimum spectral screening resolution in the 475nm≤λ≤625nm wavelength range is better than 5nm. Spectral calibration light source is white LED, and the spectral analysis was performed by a fiber optic spectrometer. The spectral calibration results were tested using a low-pressure mercury lamp, and the experimental results verified the accuracy of the spectral calibration results. The spectral imaging system proposed in this paper, has the advantages of high spectral flexibility and high optical efficiency, meeting the requirements of engineering applications.
We present an alternative approach to realize structured illumination microscopy (SIM), which is capable for live cell imaging. The prototype utilizes two sets of scanning galvo mirrors, a polarization converter and a piezo-platform to generate a fast shifted, s-polarization interfered and periodic variable illumination patterns. By changing the angle of the scanning galvanometer, we can change the position of the spots at the pupil plane of the objective lens arbitrarily, making it easy to switch between widefield and total internal reflection fluorescent-SIM mode and adapting the penetration depth in the sample. Also, a twofold resolution improvement is achieved in our experiments. The prototype offers more flexibility of pattern period and illumination orientation changing than previous systems.
In this paper, we report the coupling between the localized surface plasmon resonance (LSPR) of Au-nanoparticles and surface plasmon resonance (SPR) of the Au-film. According to the conditions for SPR excitation of the classical Kretschmann-Raether structure with 50nm Au thin film, the commonly used classes of spherical Au-nanoparticle is studied and optimized. We used the finite element analysis (COMSOL Multiphysics 5.0), to simulate the coupling. The results from calculation and simulation indicate that the resonant plasmonic coupling between Au-nanoparticles and Au-film could lead to a large field enhancement and thus improve SPR. We demonstrate that the resonant plasmonic coupling could be regulated by the size of nanoparticles, the distance between nanoparticles .
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