Bio-Rad EXQuest Spot Cutter - Manual (Page 48)

• PVDF: This option is available only when the spot cutter is in membrane cutting mode, and is
used to specify that either a PVDF or Nitrocellulose membrane will be cut. When cutting blots:
– Fill microplate wells with DI water to within 1 to 2 mm of the top. In order to release the cut
membrane into a microplate well, the cutting tip must be in contact with water.
– Do not overfill the wells/tubes, as this will allow cut membrane to float between adjacent
wells.
9. Optional. Select Make multiple cuts on large objects in the "Cut Options" panel (see Section 8.4)
to harvest large amounts of protein from a single spot and deposit it into a single microplate well.
Enter the maximum number of cuts that you want to allow in the Maximum cuts/object field.
10. Select the microplate format to be used, from the "Plate Options" panel (see Section 8.5), and then
specify the well volume and well loading order. The well volume will determine the maximum number
of spots that can be placed in a single well/tube.
11. Review the "Run Info" panel (see Section 8.8) for the number of cuts selected, the number of plates
required and the required plate well volume.
12. Click Finish to finish creating the cut request list and to open the "Plates for Cut Run" dialog (see
Section 8.9). Enter the plate name and barcode/plate ID in their respective fields. The plate barcode
can be entered using a barcode scanner or typed in. To reserve wells on a microplate, click the Edit
button for the desire plate and click on individual wells to reserve them.
13. Click Done to complete the cut request list creation.
6.2 STARTING A CUT RUN
The "Analysis Set Excision Control Tool" dialog box is used to start a cut run for gels or blots contained in
the currently loaded analysis set and to monitor the run progress. To start a cut run:
1. Make sure the spot cutter is set up as described in Sections 2 and 3 and that the required cutting
head is connected to the spot cutter.
2. Prepare the gel or blot as described in Section 4.
3. Make sure that the DI water bottle is full and the waste bottle is empty at the start of the run and
click Prime Pump.
4. If cutting blots, fill three 1.5 ml capless microtubes with water to approximately 1 mm below the tube
top and then place the tubes into the three membrane tip wash ports. After each cut, the cutting tip
is cleaned as it is dipped into the tubes of DI water.
5. If desired, review the cut request list (see Figure 6-9).
ANALYSIS SET EXCISION TOOL
6-6