A versatile method for two-dimensional electrophoresis that can be performed easily, even in small routine laboratories, is described. The procedure combines a first-dimensional isoelectric focusing run in a PhastSystem with denaturing electrophoresis in a small vertical electrophoresis chamber. The described arrangement of two first-dimensional gel strips in the second dimension allows direct comparison of two related samples, eliminating most of the artifacts that usually lead to misinterpretations. The presented procedure can be conveniently adapted to the needs of each laboratory; at present, it is being extensively used to establish protein patterns of healthy individuals of different species, a prerequisite with regard to help and support diagnosis.