Precision-cut liver slices are presently used for various research objects, e.g. to study metabolism, transport, and toxicity of xenobiotics. Various incubation systems are presently employed, but a systematic comparison between these incubation systems with respect to preservation of slice function has not been performed yet. Therefore, we started a comparative study to evaluate five of these systems: the shaken flask (an Erlenmeyer in a shaking water bath), the stirred-well (24-well culture plate equipped with grids and magnetic stirrers), rocker platform (6-well culture plate with Netwell insert rocked on a platform), the roller system (dynamic organ culture rolled on an insert in a glass vial), and the 6-well shaker (6-well culture plate in a shaking water bath). The liver slices were incubated in these incubation systems for 0.5, 1.5, and 24.5 h and subsequently subjected to viability and metabolic function tests. The viability of the incubated liver slices was evaluated by: potassium content, MTT assay, energy charge, histomorphology, and LDH leakage. Their metabolic functions were studied by determination of the metabolism of lidocaine, testosterone, and antipyrine. Up to 1.5 h of incubation all five incubation systems gave similar results with respect to viability and metabolic function of the liver slices. However, after 24 h, the shaken flask, the rocker platform, and the 6-well shaker incubation systems appeared to be superior to the stirred well and the roller incubation systems.