Modes Of Operation - tecan ls series Operating Manual

2.1.2

Modes of operation

Confocal or non-confocal
Excitation wavelengths
Emission wavelengths
Dual scanning
Resolution
Optimize sensitivity
2004-11 Operating Manual for the LS Series Scanner No. 30001798 Rev. No. 1.5
The LS SERIES SCANNERS are equipped with 3 different user selectable
pinholes for selection of the appropriate depth of focus. The smaller the pinhole
the higher is the confocality of the optical system and the shorter the depth of
field. A small pinhole typically gives a better signal to background ratio, because
all background contributions from outside the focal plane, e.g. the glass material
or the backside of the slide are rejected. However for bulkier samples or samples
with an uneven or a slightly bent surface confocality can result in lower signals or
in a poor reproducibility. Here a larger pinhole is more appropriate. That's why the
LS SERIES SCANNERS offer the choice. Just choose the degree of confocality
that is best suited for the samples.
•
The LS Scanner can be equipped with up to 4lasers.
•
Every Scanner has a HeNe laser with 633nm.
•
Lasers with the wavelengths 488nm, 532nm and 694 nm are optional.
Filters are located in special filter slides. The standard configuration comes with
one filter for each excitation wavelength or laser respectively.
Additional filters can be added into the filter slides. The filter slides can be
exchanged by the user. The 'identity' of each filter slide is automatically detected
by the instrument when it is inserted into the slot.
In order to increase the overall scan speed, two colors can be measured
simultaneously with a dual channel instrument. A calibration algorithm takes care
of the crosstalk between the two colors and corrects for it online. If you have
purchased a single channel instrument, or if the wavelengths are too close to
each other, the instrument can measure the two colors sequentially as well.
The standard resolution is 10 µm.
For larger sample features there is the option to scan with 20 µm or 40 µm thus
both increasing scan speed and decreasing the amount of data.
Smaller structures can be better resolved with 6 µm or 4 µm pixel resolution.
Scan widths for a single path scan are 22 mm for 10 µm, 20 µm and 40 µm, 6 µm
pixels and 15 mm for 4 µm pixels.
For smaller scanning areas, there is a user selectable option to increase
sensitivity without changing the scan time.
Another way to increase sensitivity is to scan transparent slides upside down.
The reason is because surface bound molecules emit the greater part of their
fluorescence into the medium with higher refractive index (e.g. glass).
2. General
2-3