Advanced system check

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MPI-CBG LMF
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This section suggests protocols for the evaluation of high end light microscopes. The tests should be carried out in a regular manner. Although the protocols are set up for the use with [[confocal]] laser scanning microscopes (CLSM), most of them can be adopted for the use with wide-field systems and other types of light microscopes.
 
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# [[ASC Fluorescence and Brightfield Imaging|Fluorescence and Brightfield Imaging]]
Advanced System Check
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# [[ASC Field Illumination|Field Illumination]]
 
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# [[ASC Color Overlay|Color Overlay]]
== Protocol (draft) ==
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# [[ASC Point Spread Function|Point Spread Function]]
 
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# [[ASC Z-Resolution|Z-Resolution]]
[[image:Conva10xok.jpg with bar.jpg|thumb]]
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The individual protocols are optimized for PDF output via the "Print as PDF"-feature in the sidebar.
[[image:Conva10xok-1.jpg with bar.jpg|thumb]]
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[[category:Manuals|A]]
*fluorescence and bright field imaging (all systems)
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[[category:ASC]]
**purpose: test general performance of the system - basic functions work fine? + Koehler illumination, light paths ok?
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**requirements: Zeiss/Leica Convallaria demo sample
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**work flow: acquire two images in basic resolution (512x512), blue ex / green em plus transmitted light image, using 10x objective
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* field of illumination
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** purpose: check for even illumination
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** requirements: mirror
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** work flow: 80/20 mirror, 488 nm laser, pinhole 1 AU. focus on reflective side of mirror. try to obtain "japanese flag".
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*psf
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**purpose:
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**requirements:
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**work flow:
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*overlay UV/V and VIS (confocal systems with separate UV/V laser coupling)
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**purpose: check the coupling precision of the UV/V laser
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**requirements: 0.2/0.5µm fluorescent beads sample
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**work flow: open pinhole(s), use high resolution apochromatic lens (NA 1.2 or above), acquire two images (UV/V plus VIS channel) with good sampling (pixel size ~100nm)
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== PSF Macro ==
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=== install LUT ===
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* Mac: go to applications folder > fiji: ctrl+click on the Fiji(.app) icon > Show package content
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* create "luts" folder if it's not there yet
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* drag and drop the *.lut into it
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* (re)start fiji
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=== install macro ===
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* Fiji > Plugins > Macros > Install...
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* select macro txt file
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* macro gets installed to that menu for the current session
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=== load bead stack ===
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* Fiji > File > Open
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* select bead stack file
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* if necessary, split channels (Fiji > Image > Color > Split Channels)
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* if necessary, crop image to a single bead
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Latest revision as of 14:04, 1 October 2009

This section suggests protocols for the evaluation of high end light microscopes. The tests should be carried out in a regular manner. Although the protocols are set up for the use with confocal laser scanning microscopes (CLSM), most of them can be adopted for the use with wide-field systems and other types of light microscopes.

  1. Fluorescence and Brightfield Imaging
  2. Field Illumination
  3. Color Overlay
  4. Point Spread Function
  5. Z-Resolution

The individual protocols are optimized for PDF output via the "Print as PDF"-feature in the sidebar.

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