Pinhole , confocal scanning-beam laser

Fig. 3. Simplified diagram of the confocal components of the AC AS 570. The laser beam is expanded to provide a diffraction limited spot at the sample. A focusing lens in front of the detection system directs the fluorescence from the focal plane through an adjustable pinhole. In the bilateral laser scanning confocal microscope, a similar technique is employed with a cursor rather than a point for illumination and detection.

Figure 12. Schematic diagram of the scanning laser confocal microscope. The out-of-focus information that normally reaches the eyepiece (detector) and leads to difficulties in interpreting optical images is rejected because the optical path does not take it through the pinhole. By scanning the incident laser beam across the sample, a digitized image is constructed from the infocus light rays that pass through the pinhole.

Figure 11.9 Schematic illustration of the principles of confocal fluorescence microscopy singlephoton exdted fluorescence occurs along the entire beam path, however the insertion of a pinhole in the focal plane of the objective ensures that fluorescence from outside the focal point of the laser (dashed line) is prevented from reaching the detector. The laser is scanned in theX-K plane to build up an image of the sample for a given depth. Translation of the sample in the Z-direction allows images (optical sections) to be recorded as a function of depth...

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