Bio-Rad EXQuest Spot Cutter User Manual
Page 148
Detecting and Editing Spots
4-27
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Gaussian fit. PDQuest determines how well the spot fits the Gaussian model and
assigns a value based on this fit.
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X streaking. The software determines how much of the spot is affected by gel
streaking in the x direction.
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Y streaking. The software determines how much of the spot is affected by gel
streaking in the y direction.
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Overlap. The software determines the percentage of the spot that overlaps with
another spot or spots.
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Linear range of scanner. PDQuest determines whether the peak intensity value of
the spot is within the linear range of the scanner.
Each spot attribute is evaluated and weighted to produce a single numerical value for
Spot Quality. If a spot fits the Gaussian model perfectly, has no streaking in the x or y
direction, does not overlap with any other spot, and has a peak intensity within the
linear range of the scanner, PDQuest will assign a value of 100 to the spot.
To display the quality of all the spots in a single image, select Show Quality from the
Spots menu and click the image. To display the quality of all the spots in all the
subwindows of a selected window, select Spots > Show All Quality.
Note:
Spots of low quality may be more accurately quantitated using the spot boundary
tools (section 4.6, Spot Boundary Tools). Spots created using the boundary tools
are not assigned a quality value.
Low Quality Spots
You can identify a set of spots that do not meet a threshold quality level. You can then
flag those spots as low quality and/or redefine them using the spot boundary tools
(section 4.6, Spot Boundary Tools).
From the Spots > Low Quality Spots submenu, select Define Low Qual. Set, then
click in the MatchSet or scanset Gaussian image. In the pop-up box, enter a threshold
quality value. All spots below this threshold will be flagged as low quality.
When you click OK, the low quality spots will appear highlighted in the image, and
the number of low quality spots will be displayed at the bottom of the image window.
To re-highlight the low quality spots in the set, select Spots > Low Quality Spots >
Mark Low Qual. Spots and click in the image.