Bio-Rad PDQuest 2-D Analysis Software - Manual (Page 39)

Fig. 35. Example PDQuest Basic Excision Tool software with SYPRO Ruby stained 2-D gel.
Follow these steps when imaging fluorescent stained gel or membrane blot:
1. Check that the light toggle switch is in UV lights position.
2. Check that the aperture is open, approximately f-stop 1.4.
3. Place the fluorescent stained gel or membrane blot onto a ProteomeWorks Plus Gel Sheet
(catalog number 165-7057) and place both on the Cutting Mat on the platform of the spot
cutter.
4. Place the Gel holding bars on the edges of the gel.
5. Add a thin layer of water to the gel or membrane blot surface.
6. Close the door.
7. Select the "UV on (Fluor)" option in the "Acquire Image" section of the Basic Excision Tool
screen.
8. Enter an exposure time, in seconds (1-30 seconds is typical)
9. Click "Acquire Image". The screen will show a processing box during image acquisition.
The amber light on the side of the Enclosure labeled, "UV lights in operation" will light
during UV illumination. There is a 15 second UV lights warm up step, then the image is
acquired, and then a dark image is acquired.
10. When the image has been acquired, it will fill in the image area of the screen. The image is
automatically inverted by the software to display dark spots with a light background. If the
original fluorescent image with light spots and dark background is desired, use the
transform button to "invert image".
11. If the image does not detect enough spots or is too faint, use the transform function to
optimize the contrast in the area of interest, or increase the exposure time and re-image
the gel or membrane.
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