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In a single photon case, you can see excitation both above and below the focal plane.
One is that there is little tissue damage to the regions above and below the focal plane that are not being imaged.
This bleaches other positions of the sample above and below the focal plane as well.
Cells appearing above and below the focal plane were omitted to prevent counting cell caps [54], [55].
Confocal microscopy analysis shows that vSMC extend filopodia that co-localize with multiple endothelial cells at various points both above and below the focal plane (Figure 2D).
Sequential images were recombined to create a single image incorporating all of the endothelial cell or smooth muscle cell labeling, above or below the focal plane of the internal elastic lamina (IEL), respectively.
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Measuring 20 fibrils, an average angle of 46.2° (standard error 3.1°) was determined at the focal plane 100 nm below the cell surface, while 20 fibrils 300 nm deeper inside the cell wall are arranged at n average angle of 92° (standard error 1.8°).
According to Hendler's model, the photoreceptors of O. wendtii would have to be present in a nerve bundle located on the focal plane of the microlenses below the dorsal arm plates [ 12, 31], and would therefore be clearly aboral and distinct from the oral radial nerve [ 116].
In addition, excitation (and hence photobleaching and photodamage) is largely confined to the focal plane, whereas regions above and below experience only the relatively innocuous infrared radiation.
The focal plane was adjusted to 500 µm below the cortical surface using a tandem-lens macroscope arrangement [18].
After obtaining an image of the cortical vasculature and extent of the craniotomy under green illumination, the focal plane was lowered to 450 μm below the pial surface.
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