Hepatic regeneration from toxic or surgical injury to the adult mammalian liver, endorses different cellular responses within the hepatic lineage. The molecular mechanisms determining commitment of a cell population at a specific lineage level to participate in liver repair as well as the fate of its progeny in the hostile environment created by the injury are not well defined. Based on the role of the Notch/Delta/Jagged system in cell fate specification and recent reports linking Notch signaling with normal bile duct formation in mouse and human liver, we examined the expression of Notch1, Notch2, Notch3, Delta1, Delta3, Jagged1, and Jagged2, and delta-like protein/preadipocyte factor 1/ fetal antigen 1 (dlk) in four well-defined experimental rat models of liver injury and regeneration. Although Delta3 and Jagged2 were undetectable by reverse transcriptase-polymerase chain reaction and Northern blot, we observed the most significant up-regulation of all other transcripts in the 2-acetylaminofluorene-70% hepatectomy (AAF/PHx) model, in which liver mass is restored by proliferation and differentiation of transit-amplifying ductular (oval) cells. The most profound change was observed for dlk. Accordingly, immunohistochemical analyses in the AAF/PHx model showed a specific expression of dlk in atypical ductular structures composed of oval cells. Delta-like protein was not observed in proliferating hepatocytes or bile duct cells after partial hepatectomy or ligation of the common bile duct whereas clusters of dlk immunoreactive oval cells were found in both the retrorsine and the AAF/PHx models. Finally, we used dlk to isolate α-fetoprotein-positive cells from fetal and adult regenerating rat liver by a novel antibody panning technique.