SystemsFast scanning TIRF

Fast scanning TIRF

True iLAS2 alternative: StarSCAN & XYZebra

Observe the localization and dynamics of
molecules and processes
in an optical section near the plasma membrane

Total Internal Reflection Fluorescence (TIRF) Microscopy is based on the total internal reflection phenomenon that occurs when light passes from a high-refractive medium (e.g., glass) into a low-refractive medium (e.g., cell, water). The evanescent field produced by total internally reflected light excites the fluorescent molecules at the cell-substrate interface and is accompanied by minimal exposure of the remaining cell volume.  The power of TIRF microscopy  lies on an extremely thin axial region of fluorescence excitation  (“optical sectioning”)   that allows imaging with very low background, minimal out-of-focus fluorescence and reduced photobleaching.


A schematic of a TIRF setup showing selective excitation of fluorophores in a tissue culture cell (refractive index of 1.33 to 1.37) resting on the surface of a glass slide (refractive index of 1.518). Wavefronts from a laser excitation source pass through the glass and are reflected from the glass-buffer boundary at a critical angle, q(c), establishing an evanescent wave that travels approximately 100 nm into the cell. Fluorophores in the cell near the glass interface (shown here as small green spheres) are excited by the evanescent wave and subsequently emit fluorescence (depicted here in red), while those farther from the interface are
not excited.

The StarSCAN system is a unique multi-application device that perform real-time control of laser illumination. StarSCAN use a pair of galvanometer mirrors to spin the laser spot around a circle at the outer edge of the objective’s back focal plane. Consequently, a collimated beam with fixed polar angle and spinning azimuthal angle illuminates the specimen. The different shadowing and interference patterns at each angle thus average out during the single-frame exposure time of the camera to produce a substantially uniform illumination field. In addition, the simplicity and ultimate flexibility of XYZebra MetaMorph user program fulfilled completely the researcher’s application needs. This software module allows to easily choose between 360° TIRF (spinning TIRF) or an optimized azimuthal angle of the illumination beam (spot TIRF) to get the best TIRF illumination condition and the best quality image.

Easy to implement on any previously-installed hardware platforms (high-end video-microscope, spinning disk set up, and more).

 Motorized control of TIRF angle

 
Azimuthal Spinning TIRF
Very fast scanning of 360° azimuthal incidence angles | Elimination of interference fringes with the obtention of a spatially and temporally uniform field of illumination (azimuth averaging)

 Multi-wavelengths TIRF with independant control (6 wavelengths )

 Option: Simultaneous matching of penetration depth at multiple wavelengths

 Uniform wide-field illumination oblique illumination sectioning: HILO (Highly Inclined and Laminated Optical)

 Powered by MetaMorph XYZebra user program

 TIRF / Single Molecule Localization Techniques  (PALM/STORM)  capabilities

 Software module very simple to use with any microscopic functionality:
Easy to switch between stationary spot TIRF, spinning-spot TIRF, spot FRAP, SPIM, and more, with one or several wavelengths | Easiness of use, efficiency, ability to concentrate on the application not the instrument
 Seamless integration in MetaMorph |Time saving no need to learn a new software
 Synergy with MDA power and robustness | Reliable high-quality performance – Efficiency
 Macro capabilities | More Possibilities – easy adaptation to your present and future needs
 Perfect combination with StarSCAN module

Could be enabled on previously-installed hardware platforms (high-end video-microscope, spinning disk set up …)

Cellular dysnamic of:
 Small random motions of organelles toward or away from the membrane
 Submembrane events: exocytosis, cytoskeletal dynamics
 Submicroscopic membrane folding and indentation
 Kinetic rates of association/dissociation at the membrane
Surface biochemistry:
 Single molecule fluorescence
 Chemical kinetics and diffusion at surfaces

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