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docs:mm_dispim_plugin_user_guide [2020/04/28 19:41] Jon Daniels [Default Timing] |
docs:mm_dispim_plugin_user_guide [2020/10/02 00:50] Jon Daniels [Setting piezo/slice calibration] |
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- Click the green-bordered **Use these!** button to compute the slope and offset of the calibration relationship. | - Click the green-bordered **Use these!** button to compute the slope and offset of the calibration relationship. | ||
- Check the computed calibration using the up and down arrows in the upper right of the Setup tab (you may need to increase the step size). | - Check the computed calibration using the up and down arrows in the upper right of the Setup tab (you may need to increase the step size). | ||
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+ | For stage-scanning, | ||
Generally the calibration slope will remain relatively constant but the offset can change slightly. It is easy to update the offset without changing the slope (e.g. when introducing a new sample): | Generally the calibration slope will remain relatively constant but the offset can change slightly. It is easy to update the offset without changing the slope (e.g. when introducing a new sample): | ||
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Distinguishing two nearby point sources of light is a classic problem in optics. If a camera’s pixels are so large that both points are read by the same pixel, you won't be able to tell them apart, regardless of how good the optics. The camera resolution must be at least twice the optical resolution to avoid being the limiting factor; essentially, | Distinguishing two nearby point sources of light is a classic problem in optics. If a camera’s pixels are so large that both points are read by the same pixel, you won't be able to tell them apart, regardless of how good the optics. The camera resolution must be at least twice the optical resolution to avoid being the limiting factor; essentially, | ||
- | The standard formula for the optical lateral resolution is the Rayleigh criterion, a distance given by the formula: 0.61*lambda/ | + | |
- | Normally, for axial resolution limited by optics, the Z-axis step must be smaller than twice (per Nyquist) the optical axial resolution. Optical axial resolution, or depth of field, is usually taken to be lambda/ | + | The standard formula for the optical lateral resolution is the Rayleigh criterion, a distance given by the formula: 0.61*lambda/ |
+ | Normally, for axial resolution limited by optics, the Z-axis step must be smaller than twice (per Nyquist) the optical axial resolution. Optical axial resolution, or depth of field, is usually taken to be 2*lambda*RI/(NA^2); for the 40x 0.8 NA water objective at 500 nm, it is ~2.1 um. Therefore, the Z-step should be < 1 um. | ||
For diSPIM, we have two views that can be merged computationally. The axial perspective from each objective is a lateral perspective (with higher resolution) from the other, so we can undersample in Z to a certain extent, which is advantageous from a speed perspective. However, giving up too much axial resolution the registration of the two views will suffer; in an extreme case your Z-step could be large enough to completely skip over a point source. For this reason, we recommend the Z-step be at least as small as the objective' | For diSPIM, we have two views that can be merged computationally. The axial perspective from each objective is a lateral perspective (with higher resolution) from the other, so we can undersample in Z to a certain extent, which is advantageous from a speed perspective. However, giving up too much axial resolution the registration of the two views will suffer; in an extreme case your Z-step could be large enough to completely skip over a point source. For this reason, we recommend the Z-step be at least as small as the objective' | ||
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