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Rolly Wiegand – CALM Leica SP5 manual
Contents
1.
Starting up the SP5................................................................................................... 1
2.
Operation of the DMI6000 inverted microscope base .............................................. 3
3.
Rules for the use of the mercury vapour arc lamp.................................................... 6
4.
Image acquisition via the LAS AF software .............................................................. 8
4.1.
Switching the lasers on............................................................................................. 8
4.2.
Setting the USB control panel................................................................................... 9
4.3.
Defining the beam path settings ............................................................................. 10
4.4.
Generating new beam path settings ....................................................................... 12
4.4.1. Excitation laser line selection.................................................................................. 12
4.4.2. Spectrum view tool ................................................................................................. 12
4.4.3. Spectroscopic separation of the emitted fluorescence signal................................. 12
4.4.4. Setting the slits for the spectral separation of the emitted light .............................. 13
4.4.5. Adjusting the PMT settings ..................................................................................... 14
4.4.6. Additional transmitted light channel ........................................................................ 16
4.4.7. Saving and loading of beam path settings.............................................................. 17
4.5.
Scanning mode....................................................................................................... 18
4.5.1. Simultaneous channel acquisition .......................................................................... 18
4.5.2. Sequential channel acquisition ............................................................................... 18
4.6.
Setting the acquisition mode................................................................................... 20
4.7.
Setting the scanning parameters ............................................................................ 20
4.7.1. Image resolution ..................................................................................................... 20
4.7.2. Pinhole size ............................................................................................................ 21
4.7.3. Scan speed............................................................................................................. 21
4.7.4. Zoom....................................................................................................................... 21
4.7.5. Averaging................................................................................................................ 22
4.7.6. Accumulation .......................................................................................................... 22
4.7.7. Scan field ................................................................................................................ 22
4.8.
Acquisition of Z-stacks............................................................................................ 22
4.9.
Using the ROI tool .................................................................................................. 24
4.10. Optimising settings via the USB control panel........................................................ 25
5.
Saving and exporting data files............................................................................... 26
6.
Post-acquisition data handling and analysis........................................................... 27
7.
Shut-down procedure ............................................................................................. 28
8.
Trouble shooting ..................................................................................................... 29
