_____________________________________________________________________________________
Rolly Wiegand – CALM Leica SP5 manual
5
Fig. 6
Left side of DMI6000 stand
2.7. Light intensity adjustment
In both modes the illumination intensity can be adjusted via the top two buttons designated
as INT (intensity). The upper button increases, whereas the lower button decreases the
brightness. Thumb rule is to use as little light as possible, in particular in the fluorescence
mode, to protect the sample against unnecessary bleaching. Accordingly, the size of the
field illuminated by the mercury arc lamp for fluorescence excitation can be adjusted via
the two buttons designated as FD (field diaphragm). By decreasing the opening of the field
diaphragm, the illuminated field will decrease to the area chosen. The fluorescence light
path is also fitted with a shutter that allows to block illumination of the sample and thus to
protect it (see the yellow arrow in Fig. 7).
2.8. In fluorescence mode, three filter cubes are provided by the system with the following
properties:
Filter set commonly used for
Excitation filter Dichroic filter
Emission filter
I3
FITC, AF488, GFP
BP 450-490
510
LP 515
N2.1
AF568, Cy3, dsRed
BP 515-560
580
LP 590
A
DAPI, Hoechst
BP 340-380
400
LP 425
Note: The system is not equipped with a camera and thus image acquisition in the
reflective fluorescence mode is not possible. This functionality is purely to find and assess
your specimen before image acquisition in the scan mode. It should also be mentioned
that the given filter cubes are prone to spectral cross-talk and thus care should be taken
regarding conclusions from the observed fluorescent signal.
