C.B.S. Scientific QNX-700 - Manual (Page 17)

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4. Insert blotting casettes into core making sure that
red side faces outward. See diagram 5.
5. Close doors and re-latch by pressing down on
the white latches so that assembly looks like that
shown in figure 6. If running one blot, slide white
reservoir conversion plate into the side without
the blotting cassette.
5.2 Electro- Blotting Procedure
1. Place stirring bar in bottom corral of lower res-
ervoir. Place core/blotting cassette assemblies
into lower reservoir. The anode (red) and cath-
ode (black) electrodes are color-coded on both
the core/cassette assembly and lower reservoir.
Ensure the red dot on the cassette assemblies
are on the same side as the red receptacle on the
lower reservoir.
Attach safety cover and turn on
magnetic stirrer. If you have the cooled unit, cat.
#QNC-700, precool buffers and attach external
cooler (set to the desired temperature) to the
tubing adaptors so that the inlet is on the bottom
and the outlet is on top (see fig. 9 on page 10).
2. Pour 1.7 liters of freshly prepared, chilled (4º) buf-
fer into lower buffer reservoir. Buffer will percolate
into central core.
3. Attach safety cover. The unit should look as
shown in figure 7 and is ready for power.
4. Connect the leads to the power supply, matching
the color-coded red to red and black to black as
shown in figure 8. See Section 6.2 for recom-
mended power conditions. Begin transfer by
electrophoresis.
5.3 Removing the Blot
1. Turn the power supply off and disconnect
the leads from the power supply. Remove the
safety cover from the unit, by placing thumbs on
white posts next to red & black connectors, then
pushing down while pulling up with fingers under
lid
. Do not remove safety cover by pulling up
on leads!
2. Blotting cassettes can be removed by leaving the
core in place and opening the top latches of the
core, opening the doors and lifting the cassettes
out. Unlatch the blotting cassettes and remove
blot from blotting sandwich.
SECTION 6
Running Conditions
6.1 Recommended Power for Slab Gels:
Precise electrophoresis conditions will vary according to the number and type of
gels used, buffer conditions employed, power input, and the general goal of the
experiment. Refer to reference section 6.3 for in depth discussions on practical and
theoretical approaches to protein gel electrophoresis.
6.1.1 ClearPAGE Gels
Run Voltage Starting Current Ending Current Approx. Run TIme
180VDC 90mA/gel 40mA/gel 30-75 minutes
6.1.2 General Recommendations
If running only one gel, keep the volts the same but reduce the mA’s by half. Keep in
mind that as the thickness of gel increases, the mA’s increase proportionally.
At constant voltage, the proteins will migrate at a constant rate during electropho-
resis with adequate heating appropriate for denaturing gels. Increasing the voltage/
mA (for a single gel thickness and percentage) will speed mobility but increase the
risk of overheating.
If using the cooling capbility of the QNC-700 the power input and the migration rate
can be increased. The joule heating generated by the higher power is offset by the
cooling effect of the buffer between the gels.
6 1 3 Tris-Glycine Gels
For SDS-PAGE Tris-Glycine (Laemmli) buffer systems with two 1.0mm thick gels at
room temperature use the following conditions at constant voltage:
80VDC until samples have fully entered stacking gel
120VDC @ 60mA-90mA/gel (depending on gel type) thereafter until dye is
near bottom of gel