FV1000MPE enables brighter and deeper imaging with superb resolution

3. Auto-adjustment of the beam in accordance with the excitation wavelength and objective.

With its auto beam expander, the FV1000MPE achieves auto-adjustment of the beam diameter in accordance with the objective and excitation wavelength, providing the optimal laser beam for multiphoton excitation.

Optical parameters optimised for multi-photon excitation.

Apart from improving the coating both in the IR as well as the visible light to approximately 90% transmission, the vignetting has been reduced - allowing for a more even illumination in the focal plane.  In MPE everything is about the probability of two or more photons being absorbed simultaneously - the excitation efficiency. As this depends on the density of photons a very small focus is crucial. The XLPLN25xW-MP has a numerical aperture (NA) of 1.05. The field number - which is a measure for the field of view (FOV) - for laser scanning is 18, which fits perfectly to the possibilities of the FV1000MPE, whereas the field number for fluorescence detection is much larger: 27.5. This is important as in multiphoton imaging excitation can only happen in the focus, but the fluorescence emitted from the focal area will be scattered multiple times on its way back out of the sample. The larger the FOV is, the more of these scattered fluorescence photons will be detected.

Fluorescence detectors for high-sensitivity multiphoton imaging

*Reflected light fluorescence detector.
This detector can capture all fluorescent signals without signal passing through a confocal aperture, so this allows imaging at a high level of sensitivity and minimizes light decay.

Multi-Point Stimulation Software

This software module allows the design of fast optical stimulation experiments. For one single point, several individual points or even a whole field of points (32x32) a time protocol with one or several stimulations can be set up, using all lasers attached to the system. The experiment protocol will be carried out with high temporal precision, minimizing the traveling time from on defined point to the next. The Multi-Point Stimulation software is a very powerful tool, especially in the field of electrophysiology. With its flexibility it will allow the creation of interesting experiments in various fields of research.

Compact FLIM NDD Upgrade kit for the FV1000MPE

The compact FLIM NDD upgrade kit for the Olympus FluoView FV1000MPE allows deep tissue FLIM imaging at depths beyond 100um. Using up to 4 Olympus NDD PMTs, the upgrade provides a cost effective, highly integrated and compact solution for FLIM imaging.

What is Second Harmonic Generation Imaging?

SHG is a secondary nonlinear optical phenomenon. In SHG, the energy of 2 photons entering a specimen is combined, producing energy in the form of light. That is, the wavelength of light observed is half of the incident wavelength (the frequency is doubled). An SHG signal is not produced unless molecules in the material are non-centrosymmetric (i.e. a centre of inversion symmetry is absent).The signal is linear, so a transmitted light detector is needed. In addition, SHG signal intensity is proportional to the size of the potential, so changes in membrane potential in the vicinity of lipid bilayers of cells with a regular molecular structure can also be analyzed.

A: SHG image of neurons in dissociated culture from the mouse cerebral cortex. After FM4-64 was administered to neurons from outside the cells, the cells were irradiated with a femtosecond laser at 950 nm and the SHG signal at 475 nm was detected with the transmitted light detector.
B: An enlargement (5x) of the portion in the yellow box in image A. As is apparent, spines protruding from dendrites can be observed with SHG.
C: SHG and multiphoton images have been superimposed.
Mutsuo Nuriya, PhD, Masato Yasui, MD, PhD Department of Pharmacology School of Medicine, Keio University.

M (Multiphoton) System

This dedicated multiphoton system is not equipped with a visible light laser. Simple optics optimized for multiphoton microscopy allow a smaller footprint, simple operation, and imaging deep within the sample. The system uses a gold-coated galvanometer scanning mirror.

B (Basic) System

This system is equipped with an IR laser for multiphoton imaging and single photon laser for visible light imaging. The system is designed for varied imaging including Live Cell and in vivo imaging. Using this system with the double laser combiner enables multiphoton imaging and visible light stimulation.

S (Stimulation) System

This system is equipped with an IR laser in the scanner for stimulation. In addition to general multiphoton microscopy, the system allows pinpoint light stimulation by multiphoton excitation during imaging with a visible laser.

T (Twin) System

This system synchronizes laser light stimulation and imaging with 2 independent IR lasers. Laser light stimulation is done with a multiphoton laser, allowing pinpoint stimulation of areas deep within a specimen that cannot be reached with single photon. Multiphoton stimulation can be done at a pinpoint, allowing observation of the effect a single dendritic spine has. Laser light stimulation can be performed in 3 dimensions or locally at specific sites.