Hoefer IEF100 - Manual (Page 27)

• p21
Preparing IPG Strips
Immobilized pH Gels (IPG) are ultra thin polyacrylamide gels on
plastic support films. The gels are made with a pH gradient covalently
bonded into the gel matrix. The gels are stable and reproducible. The
gels are supplied dehydrated and should be stored at temperatures
-20 °C or below.
The IPG strips have an acidic (+) end and a basic (–) end. 2D gel
images are typically shown with the acidic side on the left, and that is
the orientation of the IPG strips in the IEF100. The proper electrode
must be applied to each end: anodic (+) electrode to the acidic end
(left) and the cathodic (–) electrode to the basic end (right).
The dried strips require rehydration before the run. Typically, several
different reagents are included in the rehydration solution to help
solubilize the proteins, and enable successful isoelectric focusing. Rehy-
dration is typically done at room temperature, or 20 °C, and needs a
minimum of 8 hours for good absorption of the reswelling solution. It
is frequently more convenient to allow rehydration to occur overnight.
During rehydration, the IPG strips are covered in mineral oil to prevent
loss of moisture, and prevent urea crystallization.
The IPG strips are rehydrated using one of the rehydration/equilibra-
tion trays supplied with the IEF100. Protein sample can be added to
the IPG strip in this step. The IPG strips are transferred to the IEF100
for focusing.