Hepatocytes are the primary defining cell of the liver, performing the vast majority of its functions (Wallace et al., 2010a). Isolation and/or culture of hepatocytes are therefore common experimental techniques employed in the study of liver functions (Wallace et al., 2010a). Several factors limit hepatocyte utility. Hepatocytes do not proliferate in vitro and therefore cannot be expanded in vitro. Furthermore, culture results in dedifferentiation and loss of function. Complex culture modifications can ameliorate this loss but experimentally introduce an array of often uncharacterized factors, which can complicate interpretation (Wallace et al., 2010a). In many cases, human hepatocytes would be the ideal species to use for most experiments related to toxicity. However, human liver is in short supply and is often of poor quality. In the absence of sufficient human hepatocytes from donor livers, the main alternative for generating human hepatocytes is through differentiation of human embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) (Lavon et al., 2004; Rashid et al., 2010; Takayama et al., 2012; Zhang et al., 2013). Despite significant progress over the last 12 years, it has not been possible to generate hepatocytes with function quantitatively similar to adult human hepatocytes (Takayama et al., 2012). Stem cell–derived hepatocytes remain in a fetal state and cannot progress further unless transplanted in vivo (Lavon et al., 2004; Rashid et al., 2010; Takayama et al., 2012; Yu et al., 2012; Zhang et al., 2013). Because normal hepatocytes dedifferentiate into a fetal state in vitro (even when present within culture tissue slices) (Wallace et al., 2010a; Wright and Paine, 1992), it may be not be surprising that this barrier exists in stem cell–derived hepatocyte culture. In addition, a major hurdle to the use of ESC/iPSC-derived hepatocyte is their high cost of generation. As part of an European Commission–funded project, the cost of hepatocytes derived from iPSCs was calculated, taking into consideration the most recent research that compared drug metabolism activity in iPSC-derived hepatocytes with human hepatocytes (Takayama et al., 2012). In order to obtain the hepatocytes, the iPSCs required a 4-stage differentiation protocol with a variety of recombinant growth factors. In addition, the cells were infected with adenovirus directed to overexpress 2 transcription factors. Considering growth factors alone (without taking into account the cost of culture media, virus production, culture ware, and the cost of failures), it was calculated that the cost of generating hepatocytes is approximately 5 million times greater than using glucocorticoid and a human cell line with B-13 properties (d-LIVER consortium, 2012). With current technologies, it is unlikely that ESC- or iPSC-derived hepatocytes will be a practical solution in routine in vitro toxicity testing. The B-13 cell could offer a potential route to delivering a cost-effective, simple solution to the production of functional hepatocytes in vitro. This rat pancreatic acinar-like cell line is readily expandable in simple culture medium and in response to one simple glucocorticoid hormone differentiates into nonproliferative hepatocyte-like (B-13/H) cells (Shen et al., 2000; Wallace et al., 2010b). B-13 cells model a pathophysiological response of rodent and human acinar tissue to differentiate into hepatocytes both in vitro and in vivo on exposure to high levels of glucocorticoid (Fairhall et al., 2013b; Wallace et al., 2009, 2010c). The cells retain a degree of biological stability in that they have maintained a normal karyotype in terms of the number of chromosomes per cell (although with some cytogenetic abnormalities); do not grow in soft agar (in contrast to tumorigenic cells); and are able to selectively engraft into the pancreas and liver (Fairhall et al., 2013a). Within the liver, B-13 cells differentiate into hepatocytes (Fairhall et al., 2013a). The intrinsic value of this cell line is that it offers an unlimited and reproducible supply of hepatocytes in vitro, without the requirement for tissue donors. Given both the simplicity and costs of a B-13 approach to hepatocyte generation, it is likely that a human B-13 equivalent would have utility in both experimental studies (eg, toxicity screening) and clinical applications (eg, extracorporeal liver support). In this paper, we demonstrate that B-13/H cells recapitulate the sexually dimorphic expression of cytochrome P450 (CYP)2C11; retain CYP1-3 family induction to classic inducers; and express variable levels of functional transporter expression. Stable transfection of the human CYP1A2 gene resulted in a transgenic cell line with active human protein that metabolized a probe substrate and the progenotoxin 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (PhIP) to a genotoxic product.