CCMI Leica SP2 AOBS DIC Quick Ref

Establish Kohler illumination:

Set scope to USER (white light).
Ensure Vis/Scan mirror switch on condenser turret is set to Vis.
Adjust white light intensity to preference.
Focus on specimen (you may need to switch back to UV to do this).
Close condenser field iris (move upper lever to left).
Open condenser aperture iris (move lower lever to right).
Focus image of field iris with condenser focus control.
Open field iris until image is almost the same size as your field of view.
Center image of field iris with condenser centering controls.
Open field iris until image moves out of field of view.
Close aperture iris about half way.

Ensure upper and lower polarizing filters are inserted.
Make fine adjustments to the upper polarizer to get darkest image.
Set prism on condenser turret (upper) to match the objective in use.

NB: 20x/0.7 IMM uses the 40x prism.

Set prism on objective turret (lower) to C, D or E depending on objective in use.

20x/0.7 IMM and 40x/0.85 use C.
63x/1.2 Water uses D.
63x/1.4 Oil uses E.

Make fine adjustments to DIC prism position to optimize contrast.
Set mirror above condenser to SCAN (care - turn SLOWLY!).
Set lower polarizer to the OUT position
Activate transmitted light detector in software.
Set scan control to CONTINUOUS.
Make fine adjustments to DIC prism position, upper polarizer and detector controls for best image.

Note: For best fluorescence resolution it is best to proceed as follows:

Acquire fluorescent image with DIC prism set to BF.
Turn off fluorescence detectors and turn on transmitted detector.
Acquire DIC image with DIC prism set to C, D or E.
Overlay images in software (<Process>->Editing->Merging - select Ch, check Append box.)

jdc 0409; Last modified 041011