Understanding the hepatotoxicity of drugs and chemicals is essential for progress in the pharmaceutical industry, medical science and academic research. The study of hepatotoxicity in vitro is complicated by the difficulty of maintaining hepatocytes in culture due to a lack of understanding of the humoral and matrix requirements of these cells. A variety of in vitro models of the liver have been developed, such as perfused livers, liver slices and three-dimensional perfused bioreactors, but the static cell culture is the most commonly used system. In this review we present the advantages and disadvantages of each system and their roles in the study of hepatotoxicity. We will also discuss how the various culture conditions such as medium and matrix composition affect the systems. The technological advances, which started the fields of genomics, proteomics and metabonomics are playing a very important role in uncovering novel biochemical pathways and markers of toxicity. Several of these studies have focused on hepatotoxicity, particularly on the effects of acetaminophen, carbon tetrachloride and aflatoxin B1. Finally, we will discuss the new field of systems biology, which focuses on interpreting and integrating data from all of the other fields.