The aim of this study was to assess directly the function of isolated hepatocytes 1 year after transplantation into the spleen, using an original model of isolated rat- spleen perfusion. Three specific liver functions, albumin synthesis, indocyanine-green clearance, and antipyrine oxidation, were studied. Five ×106 isolated hepatocytes were injected into the spleen of syngenic Wistar-Furth rats. One year later, splenectomy was performed, and the splenic pedicle was carefully isolated in order to allow a selective ex vivo perfusion for 3 hr. De novo albumin synthesis was studied by qualitatively using immunoelectrophoresis and autoradiography, and quantitatively using (35S)- methionine incorporation in albumin. De novo albumin synthesis was observed in spleens containing transplanted hepatocytes but not in controls (P<0.001); (35S)-methionine incorporation was significantly higher in spleens containing transplanted hepatocytes than in controls (132±67 cpm/spleen/hr vs. 14± 6 cpm/spleen/hr, P<0.001). Antipyrine clearance was significantly higher in spleens with transplanted hepatocytes than in controls (67.4±4.9 μ1/min/g vs. 0.2±0.4 μl/min/g, P<0.0l). No statistically significant difference was observed with indocyanine-green clearance (4.2± 6.0 μl/min/g, vs. 5.2±5.1 μl/min/g, P>0.05); this was probably due to the absence of compartmentation between the sinusoid and biliary sectors in this model. In conclusion, using this original isolated rat-spleen perfusion model, it was directly observed that 1 year after transplantation, intrasplenic hepatocytes can perform two liver-specific functions, i.e., de novo albumin synthesis and antipyrine clearance. © 1990 by Williams and Wilkins.