Cultured precision-cut liver tissue slices are useful for studying the metabolism and toxicity of xenobiotics in liver. They may also be used to investigate the behavior of and interaction between different cell types in an intact histo-architecture. Because cultured liver tissues undergo a loss of function and morphology because of their separation from the blood supply, we investigated changes in key protein marker expressions in parenchymal and non-parenchymal cells, as well as in the extracellular matrix (ECM) at different time points. We also compared conventional culture methods such as static and dynamic cultures with perfusion culture, which allows a continuous exchange of the culture medium. In conventional culture methods, the expression of vimentin and collagen type IV decreased after 5 h in the non-parenchymal cells and the ECM, respectively, whereas the hepatocyte nuclear factor 4 alpha (HNF4alpha) staining in the hepatocytes remained constant. In perfusion culture, on the other hand, vimentin, collagen type IV, and HNF4alpha staining were clearly detectable after 5 h. The histo-architecture obtained from perfusion culture was also more compact than those obtained from conventional culture methods. After 24 h, only the perfusion cultured sample retained protein marker expression in all components of the liver tissue. Our results suggest that, to develop improved culture techniques for liver slices, changes at the early time-points should be taken into consideration. Our results also show that culture techniques that enable a continuous exchange of the culture medium seem to be superior to static or dynamic cultures in terms of maintaining the protein expression and the histo-architecture.