Bio-Rad Components for Older Model Spot Cutter - Manual (Page 34)

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Lights not turning on during image acquisition and cutting tip not turning
during cut
• The spot cutter and computer "power on" sequence were done in the incorrect order. The
computer needs to be on and fully booted up before turning on the spot cutter. Use the
following steps to return the spot cutter to full communication mode.
• Close PDQuest (whichever spot cutter screen you are in). Turn off the power on the spot
cutter (switch the front black I/O switch to O). Wait a few seconds. Turn on the spot cutter
(switch the front black I/O switch to I) and push the yellow "Start/Pause" button. Let the spot
cutter run through its home position routine until it returns to the back right position (park
position).
• Open PDQuest and go to either Basic Excision Tool, or Excision Gel Selection. Full
communication to all commands should be restored now.
Spot Cutting and Pickup
In the best conditions, the mechanism of cutting is very reproducible. Normal operation
produces cutting and pickup efficiency at >95% of the spots cut. For any spots not picked up
on the first cutting pass, the software allows recutting of specific spots based on microtiter plate
well number. The second cutting pass generally picks up any spots not picked up the first
time.
Spot Cut Pick Up Rate
Acrylamide gels are effectively cut with the spot cutter when they are laying on the cutting mat
or the Gel Sheet. It is also effective at cutting gels which are bonded to a plastic backing.
Varied results have been obtained using gels bonded to glass plates.
If the pickup rate is less than 95 spots out of 100 cuts, please review the wetting process, the
cutting tip depth and cleanliness, and the level of the instrument. These are the most
common sources of poor pickup rates.
a. The wetting instructions are listed in the Gel Preparation section before this section.
b. Cutting tip depth should be adjusted so that the cutting tip is going just below the gel and
scoring into the Gel Sheet below the gel. If the tip is not going down far enough or is
going down too far, adjust the cutting tip height as described in Section 3.5 "Setting
Cutting Tip Height".
c. The cutting tip should be cleaned daily after spot cutting. This consists of taking the
cutting tip off the assembly and rinsing with water, as from a squirt bottle. If the pickup
rate on spots is decreased or less than 95% of cut spots, a more aggressive cleaning can
be used. Prepare a detergent solution. Soak the tip for a few minutes to overnight. Scrape
the inside of the tip with a metal probe (a straightened paper clip works well) to dislodge
any acrylamide that may have stuck to the inside of the tip.
d. Using the Wash Wells. The wash wells may help improve the pickup rate.
e. The bubble level on the instrument is used to assess the level of the cutting platform. If
the instrument is off level, one portion of the gel may dry out more quickly than another
and can contribute to poor cutting and pickup efficiency.
If all the settings seem to be correct, contact Bio-Rad for further troubleshooting.