Bio-Rad EXQuest Spot Cutter User Manual
Page 52
Figure 6-14. Align Analysis and Cutter Images: Step 2 – Perform Alignment Panel
•
Automatic Alignment. Automatic alignment uses a Gaussian fitting algorithm to align the
cutter image spots with those in the analysis set image. If the cutter image spots do not
align correctly, make sure that the file size is set correctly on the device used to scan the
analysis set image. For example, the FX, Densitometer, and the GS series automatically set
file sizes, whereas camera scanners cannot. If your analysis set image was generated from
a camera acquisition device, such as the Gel Doc
™
EQ, ChemiDoc
™
EQ, or VersaDoc
™
, it
will not have specified file dimensions. For such images, you will need to enter the physical
size dimensions correctly at the time of acquisition. It is important to have correct physical
dimensions before you proceed with the spot detection and match set creation, otherwise
auto-alignment will not work correctly.
•
Manual Alignment. Manual alignment positions cut marks at the location defined by the
green data area box and in the same relative positions that the spots appear in the analysis
set image. It does not adjust the cut positions to optimize the fit. Manual alignment should be
done when there are only a few spots to align.
c.
Set the Confidence threshold to an acceptable level. We recommend it to be a high value. Cut
marks will be colored green for spots that have an alignment confidence greater than the
Confidence threshold or colored red for those spots that have an alignment confidence below
the confidence threshold.
Figure 6-15. Align Analysis and Cutter Images: Step 3 – Confirm Alignment Panel
d.
Click Jump and confirm to visit each spot with an alignment confidence level less than the
Confidence threshold. If a spot is not correctly aligned, click and drag it until it is properly
aligned. Note that the cursor will display the confidence level as the spot is moved. Repeat the
above procedure for all spots with a low confidence level. Note that the green cut circle size
represents the actual cut size and depends on the cutting tip size selected in Step 7 of
Section 6.1.
ANALYSIS SET EXCISION TOOL
6-10