Biology Reference
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2. Store the specimen at 4 °C or proceed directly with sample
scanning via automated microscopy ( see Note 21 ).
The automated microscope system “.slide” allows digitalizing a
large number of slides with minimal human effort during the scan-
ning process (a video recording showing this procedure can be
downloaded from the authors' website at http://www.ceitec.eu/
functional-genomics-and-proteomics-of-plants/rg46#vybaveni ) .
3.5 Automated
Microscopy Procedure
1. Before the scanning is initiated, up to 50 slides are loaded into
the slide loader. The robotic arm scans the slides via bar code
reader ( see Note 22 ). If the slides are not designated by bar
codes (or the codes are not detected), the reader mechanically
detects the presence of a slide in the respective loader position.
After all the slides in the slide holder are successfully detected,
the scanning process can be initiated.
2. The next step is to set up several scanning parameters: (a)
information and designation as to the type of scanned slide ( see
Note 23 ), (b) slide positions in the loader that will be scanned
(in most cases, all slides are selected), (c) magnifi cation at
which the slides will be scanned ( see Note 24 ), (d) position of
image storage area on disk, (e) acquisition properties ( see Note
25 ), (f) specifi cation of the slide scanning area ( see Note 26 ),
and (g) optionally, in a case of diffi cult specimens, image qual-
ity can be improved by using the extended focal imaging (EFI)
scanning mode ( see Note 27 ).
3. The scanning process can now be started simply by pushing
the START button.
4. Slides are subsequently loaded from the slide loader by robotic
arm (via a vacuum-based adaptor) and placed into the motor-
ized stage ( see Notes 8 and 20 ).
5. As the fi rst step, the system creates an overview of the slide at
low magnifi cation (2×). In the slide overview, contrasting
objects are automatically recognized. Slides are scanned
according to the user settings ( see step 2 ).
6. If the manual adjusting mode of the scanning area was selected
( see step 2 , f), then the slide scanning area must now be set.
This scanning area will be used for all slides in the loader that
are selected to be processed.
7. In the next step, a focusing map is automatically set up ( see
Note 28 ) and the scanning process initiates. The fi nal scanned
image is composed of small overlaid frames, each correspond-
ing to the fi eld of view of the respective objective. The image
processing and creation of the fi nal image (virtual slide) runs
in real time and can be observed on the computer screen.
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