Excitation Angle Tab - tecan ls series Operating Manual

6.3.7

Excitation Angle Tab

Default
2004-11 Operating Manual for the LS Series Scanner No. 30001798 Rev. No. 1.5
The excitation beam of the laser scanner usually strikes the sample with an angle
of approximately 25° (see Figure 6-10). This improves the confocality and
minimizes unwanted stray light effects due to back reflections. Therefore this
configuration (='Default') gives the best performance for standard slides and
other flat sample formats.
However there might be applications where the default angle of incidence is not
optimal or the samples cannot be measured at all. A typical example is a
microplate with printed arrays at the bottom of the wells. When measured from
the top an inclined laser beam would usually be blocked by the walls and would
not reach the bottom of the well. The variable beamshift feature of the laser
scanner allows these samples to be measured as well. The angle of incidence for
the fluorescence excitation can be varied between 0° and 25° in steps of 0.1° via
the software.
The geometry of the sample determines the angle of incidence. The angle should
be set to a value that allows the excitation beam to excite the full area of interest
at the bottom of your sample without hitting the wall of the MTP before reaching
the measurement surface.
If the angle for the scan is changed, the angle used for autofocus should also be
adjusted. Because the autofocus does not work with normal incidence, the lowest
angle accepted by the scanner is 10°. If your sample has high walls we
recommend using the lowest possible value (=10°).
If 'Wellmode' is selected (in the advanced autofocus settings menu), except for
the image scan, it is not necessary that the full area covered with spots is hit by
the laser beam for the autofocus. In that case some shadowing effect can be
tolerated. A minimum of 2.0 mm at the center part of the bottom is required
however.
It is recommended to select Default for all 'flat' samples (e.g. slides).
Angle is set to 25° for both the image scan and for autofocus.
6. LSServer, Scanner Shell and Download
6-49