In fact, confocal technology is proving to be one of the most important advances ever achieved in optical microscopy. In a conventional widefield optical epi-fluorescence microscope, secondary fluorescence emitted by the specimen often occurs through the excited volume and obscures resolution of features that lie in the objective focal plane. The problem is compounded by thicker specimens (greater than 2 micrometers), which usually exhibit such a high degree of fluorescence emission that most of the fine detail is lost. Confocal microscopy provides only a marginal improvement in both axial ( z along the optical axis) and lateral ( x and y in the specimen plane) optical resolution, but is able to exclude secondary fluorescence in areas removed from the focal plane from resulting images. Even though resolution is somewhat enhanced with confocal microscopy over conventional widefield techniques, it is still considerably less than that of the transmission electron microscope. In this regard, confocal microscopy can be considered a bridge between these two classical methodologies. Presented in Figure 1 are a series of images that compare selected viewfields in traditional widefield and laser scanning confocal fluorescence microscopy. Digital optical converter madison wi series# A thick section of fluorescently stained human medulla in widefield fluorescence exhibits a large amount of glare from fluorescent structures above and below the focal plane (Figure 1(a)). When imaged with a laser scanning confocal microscope (Figure 1(d)), the medulla thick section reveals a significant degree of structural detail. Likewise, widefield fluorescence imaging of whole rabbit muscle fibers stained with fluorescein produce blurred images (Figure 1(b)) lacking in detail, while the same specimen field (Figure 1(e)) reveals a highly striated topography in confocal microscopy. Digital optical converter madison wi series#. Digital optical converter madison wi serial#.
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