Imaging Core
Equipment

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Leica SP8 Confocal Microscope

Equipment Location: 
Basement, Room B04
Equipment Manager: 
Cyril Eleftheriou, Ph.D.
Leica SP8 Confocal Microscope

Advantages

Confocal microscopy enables acquisition of 2D or 3D fluorescence images as well as time-lapse images by collecting emission light only from the focus of a laser beam. Excitation light generates fluorescence throughout the specimen and out-of-focus light is eliminated by a pinhole. This microscopy can be used for imaging multi labeled structures with submicron resolution in live or fixed tissue. The best quality is achieved close to the surface of the sample (<200 μm). However, with the advancement of tissue processing, large cleared pieces could be imaged (up the entire mouse brain or spinal cord).

Description

Leica SP8 upright confocal microscope is equipped with Lightning SuperResolution module and scanning DM6 CS stage. Transmitted light pictures are combined with fluorescent images in the same software. The setup combines four lasers (405 nm, 488 nm, 552 nm, and 638 nm) and three filter-free spectral detectors (2 HyD and 1 PMT) for up to five individually regulated channels. Switchable between resonant and nonresonant mode for optimal scanning speeds. Using resonant scanning a higher fluorescence quantum yield and reduced photobleaching are achieved. The continuously adjustable speed of the nonresonant scanner (1-1800 Hz) is ready for single molecule imaging, image correlation and imaging of rapid dynamics alike (up to 72 frames / second at 512 x 32 scan format). The maximal image resolution of 64 Megapixels allows sampling optimally, also with low magnification lenses. Structures up to 120 nm in several colors can be resolved simultaneously and at high recording speeds. This microscope can do tiling.

The microscope has the following objectives: 5x (air, WD 13.0, NA 0.15), 10x ( air, WD 2.2, NA 0.4), 10x (air, WD 11.0, NA 0.3), 20x (water, WD 0.62, NA 0.75), 40x (water, WD 3.3, NA, 0.8), 63x (oil, WD 0.14 NA 1.4). Multiple digital zoom options are available.

Data analysis is typically conducted on a different computer using Fiji or Imaris.

After individual consultation and training sessions, researchers are given access to schedule instrument time through the institutional online booking system “booked”. Assistance with imaging and experimental design is available during normal business hours.