Core Facility - Confocal digital analysis

Project Summary

Confocal laser scanning microscopy (CLS) is a digitized and high-resolution imaging method based on fluorescent signals derived from the focal plane and photo multiplier (PMT) detection in fixed biological samples. In principal, the signal „noise“ present in light- and epifluorescence microscopy due to focus variables and thickness of samples (e.g. known as diffuse signals) can be excluded in confocal imaging by pinhole exclusion (5-180µm) point-to-point scanning and optical dissection at XY or XZ-axis (i.e. image stacks) leading to high-resolution confocal images of subcellular structures (similar to a „cell-CT“). Fixed cells or tissue sections stained with 488-633nm fluoroprobes (cell markers, antibodies etc.) can be investigated using either single, double or triple labeling techniques. Our LEICA CLS device consists of a standard (DM-RE) and inverse microscope stage (DM-IRBE) and a three channel multi-laser device with an Ar-Laser (488nm, blue), HeNe-laser (543nm, green) and HeNe-laser (633nm, red) including image software pack (Leica Microsystems Inc., Bensheim, Germany). The device does not include a two-photon-laser module for e.g. live cell recordings.

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