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CHAPTER 2 - SAMPLE PREPARATION
Lightsheet Z.1
Tips, Troubleshooting and Additional Information
Carl Zeiss
02/2013
000000-1790-528
37
4
Tips, Troubleshooting and Additional Information
4.1
Tips
Protocols
As a general rule, follow the protocol carefully and try it out well in advance as you may encounter
difficulties and missing parts (chemicals…) that may hinder you to use the sample straight away. The
protocols presented in this section are not the ultimate solution for every problem. They can be improved
and adapted to your need. If you encounter a problem, check the scientific literature and the other
protocols presented and see if you can find a tip that you may apply to solve your problem. Be creative.
Safety
Please refer to the safety instructions provided with the Lightsheet Z.1.
Setup Requirements and Maintenance
For optimal performance of the Lightsheet Z.1, please check the Setup Requirements information
delivered with the system.
For information about maintenance and cleaning any part of the Lightsheet Z.1, including the optics,
please refer to the according
CHAPTER
in this manual.
Refractive index mismatch
Light is refracted when it crosses the interface between two media of differing refractive indices (RI).
Mismatching the refractive index of the objective immersion medium and mounting medium is one of the
main causes of image degradation in microscopy. Refractive index mismatch results in
stretching/compression of the z-axis. Also, spherical aberration is worsened by axial spreading of the
point-spread function (PSF) resulting in reduced axial resolution. This phenomenon is exacerbated with
depth and with a high numerical aperture objective, serious problems arise when imaging deeper than 10
µm into an aqueous sample. The mounting medium and the immersion medium should be matched. It is
not a major issue in LSFM but this as to be considered when filling the imaging chamber in regard to the
sample preparation technique, especially for embedding as gelling agents are used.
4.2
Troubleshooting
Common problems
LSFM is a fluorescence microscopy technique so many troubleshooting guidelines from other microscopy
techniques apply here as well. Do not hesitate to ask experts in the field, check the literature, as well as
internet resources that may provide you with a more detailed solution to the problem you have
encountered. Also check with your Lightsheet Z.1 specialist for FAQs and tips for troubleshooting.
Содержание Lightsheet Z.1
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