BIO RAD PROTEAN II xi CELL Setup Manual

PROTEAN
This kit allows 18.5 cm wide x 20 cm long gels to be run on Bio-Rad's PROTEAN II xi Cell thus accommodating commercially
available immobilized pH gradient gel strips for 2-D second dimension protein separations. It contains:
• Wide sealing gaskets for the upper buffer chamber (2)
• 20 cm glass plates (2 sets)
• 20 cm notched clamps (2 sets)
• Foam gaskets for the PROTEAN II xi Casting Stand (2)
• Alignment card
• Narrow spacers (4) *
• Prep Combs (2)*
* 1 mm or 2 mm thick depending upon which kit you ordered.
Casting Gels
Gels are cast the same way as with the standard PROTEAN II xi Cell (refer to your user manual for details). For best results, replace
the solid rubber gaskets on the PROTEAN II xi Casting Stand with the foam rubber gaskets included with this kit. They provide a
more reliable seal when using the narrow spacers. When assembling the gel "sandwich", use the alignment card to push the spac-
ers to the extreme outer edges of the glass plates to ensure maximum gel width. Since the card is narrower than the gel cavity, you
will need to slide it side-to-side to properly align the spacers.
After pouring the monomer solution, insert the comb into the gel. Using combs ensures gel-to-gel dimensional repeatability. This
is especially true if using the PROTEAN II xi Multi-Gel Casting Chamber. Overlaying gels with t-amyl alcohol can be done,
but unless it is applied simultaneously to all gels, the gel heights will not be the same because the weight of the overlay applied to
the first gel (and each subsequent gel) will displace the solutions in the other gels.
The combs provided in this kit have two sides—one with a reference well for standards and one without. The side without the stan-
dard lane allows for commercially available 18 cm IPG strips to be used without excess trimming. The side with the end reference
well allows standards to be run along side your sample. However, 18 cm IPG's will need to be trimmed more to fit into the small-
er prep well. Deciding which comb side to use depends upon what you know about your sample. When running without the refer-
ence well, an internal standard can still be included by adding a known protein to your sample. Providing a marker between gels helps
to monitor the reproducibility of the 2-D patterns and makes post run gel comparisons easier. To determine which standard mark-
er to use, run a 1-D SDS gel of your unknown sample.
Assembling the Upper Buffer Chamber
Replace the standard U-shaped gaskets on the central cooling core with wider U-shaped sealing gaskets provided with this kit.
(Refer to section 5.1 in the PROTEAN II xi Cell manual.)
Once the gels have polymerized, remove the gel sandwich assemblies from the casting stand, remove combs, and mount the gel sand-
wich assemblies onto the central cooling core to create the upper buffer chamber.
If running one gel, a buffer dam can be mounted on the opposite side of the core. If you do not have a buffer dam, clamp an inner
and outer glass plate together without spacers and mount them on the central cooling core. Note that either U-shaped gasket size
will work with the buffer dam or glass plate set.
® II xi Cell IPG Conversion Kit
Setup Guide