Light sheet and confocal microscopy
Light sheet microscopy
Light sheet microscopy – also referred to as single plane illumination microscopy (SPIM) is a is a gentle way of imaging sensitive samples or fast biological processes in vivo. The specimen is illuminated only in a single plane at a time and detected from the perpendicular direction. Since there is no out-of-focus excitation, phototoxic effects are reduced to the focal plane. Light sheet imaging has intrinsic optical sectioning. By moving the sample through the light sheet, 3D images of a specimen can be recorded. With this technique large 3D volumes are captured at a much higher speed but slightly lower resolution compared to confocal imaging.
Leica SP8 DLS specifications
Our Leica SP8 DLS (inverted microsocpe) is equipped with:
Light source: five lasers with wavelenghts:
- 405 nm
- Solid state 488 nm
- Solid state 514 nm
- Solid state 552 nm
- Solid state 638 nm
Detectors: two PMT:s (photomultiplier tubes), one HyD GaAsP-spectral detector, one PMT for transmitted light
Objectives for confocal imaging: 10x/0.3, 20x/0.75 IMM, 25x/0.95W, 63x/1.4 Oil, 63x/1.3 GLYC
Objectives and mirrors for light sheet microscopy: imaging 25x/0.95W DLS, 10x/0.3W DLS, illumination 2,5x/0.07, 5x/0.15, DLS TwinFlect 2.5mm, DLS TwinFlect 5mm
Software: LAS X with HyVolution, Dye Finder, 3D Visualisation, Microlab (for FRAP, FLIP and FRET) and Huygens Base Package for Deconvolution
Confocal microscopy – or to be more precise: confocal laser scanning microscopy (CLSM) – is a method that is particularly suited for microscopy of fluorescent or reflecting specimens. The confocal microscope eliminates out-of-focus light, and creates a clear view of structures in the focal plane – also in specimens that are to thick for conventional fluorescence microscopy.
If you acquire a series of confocal images at different depths in a thick specimen, the image series can be used for 3D analysis and 3D rendering and you can visualise structures that could not be distinguished in conventional light microscopy.
If you are publishing an article with results obtained with the microscopes belonging to the Department of Biology, please make sure to acknowledge us:
"We acknowledge the Microscopy Facility at the Department of Biology, Lund University".