20 – LSM 880 Training Guide
Understanding Image Quality
Pinhole (Confocal Aperture)
Ideally, the confocal aperture should be set to the size of the structure(s)
you are trying to resolve. However, for example, some sub-cellular
structures of interest may be beyond Abbe’s diffraction limit and
therefore beyond the microscope’s capability to resolve. The confocal
aperture has some practical limits that can be used to guide the usage
of this setting.
Start by closing down the confocal aperture to 1 AU. You can certainly
close the confocal aperture below this value and continue to improve
resolution, just understand that below 1 AU you will lose signal level at
an exponential rate.
Conversely, you can increase the confocal aperture diameter to improve
the detected signal level if you can sacrifice axial resolution.
In some cases, the signal level may be so low that increasing the aperture diameter is the only way to lower the gain enough to
get a usable image.
In other cases, if the gain is too high (causing excess noise) and the laser power cannot be increased due to photo effects then
increasing the confocal aperture is the only option to improve image quality.