As first dimension an IEF is done. The proteins were separated according to their isoelectric points.
Because a immobilized pH-gradient is more reproducable the IPG-strips are commonly used as first dimension. These strips run denaturated with 8 M Urea so that also the non water-soluble proteins will be displayed.
The proteins within their gel-strips were equilibrated with the normal SDS-sample buffer.
Then as second dimension a SDS-electrophoresis is done. Depending of the amount of different spots the second dimension is done also in large sized gels.
The second dimension functions vertically and als horizontally.