light pathway in Confocal Microscopy - أطلس مختبرات العرب

Confocal microscopy offers several advantages over conventional optical microscopy, including shallow depth of field, elimination of out-of-focus glare, and the ability to collect serial optical sections from thick specimens. In the biomedical sciences, a major application of confocal microscopy involves imaging either fixed or living cells and tissues that have usually been labeled with one or more fluorescent probes

When fluorescent specimens are imaged using a conventional widefield optical microscope, secondary fluorescence emitted by the specimen that appears away from the region of interest often interferes with the resolution of those features that are in focus. This situation is especially problematic for specimens having a thickness greater than about 2 micrometers. The confocal imaging approach provides a marginal improvement in both axial and lateral resolution, but it is the ability of the instrument to exclude from the image the "out-of focus" flare that occurs in thick fluorescently labeled specimens, which has caused the recent explosion in popularity of the technique. Most current confocal microscopes are relatively easy to operate and have become part of the basic instrumentation of many multi-user imaging facilities. Because the resolution possible in the laser scanning confocal microscope (LSCM) is somewhat better than in the conventional widefield optical microscope, but still considerably less than that of the transmission electron microscope, it has in some ways bridged the gap between the two more commonly used techniques. Figure 1 illustrates the principal light pathways in a basic confocal microscope configuration.