Bio-Rad PDQuest 2-D Analysis Software - Manual (Page 57)

8.4 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 or glass backing. 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. For free gels, the 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 7.4
"Set 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. Using 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.
In depositing spots to the microtiter plate, it is important to prepare the plates with liquid in the
wells. The liquid can be the solution used as the first step in the digestion process, or any other
liquid used in typical lab conditions. The stainless steel cutting tip and polypropylene microtiter
plates are compatible with a wide variety of chemicals. If all the settings seem to be correct,
contact Bio-Rad for further troubleshooting.
8.5 Technical Service
For technical assistance with the system including all hardware and software, contact your local
Bio-Rad office. All spare parts not listed in this document can be ordered by
contacting your local Bio-Rad office. For inquiries and requests regarding system repair or service,
contact your local Bio-Rad office or distributor. Please have the following details available:
1. Instrument model and catalog number.
2. Serial number (located on the back of the system).
3. Hardware and software version information.
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