Galactosyltransferase has been isolated from putative preneoplastic hepatocyte nodules generated in the resistant hepatocyte model by the procedure of Solt et al. (Am. J. Pathol., 88 (1977) 595-609). The following observations have resulted from these studies: (a) the specific activity of galactosyltransferase isolated from hepatocyte nodules by affinity chromatography was reduced to about 1/3 that of the enzyme in control and in liver tissue surrounding the nodules; (b) the galactosyltransferase activity from normal rat serum eluted from the alpha-lactalbumin affinity column as a single peak (spec. act. = 1.57 nmol/min per mg) while that from the serum of nodule-bearing rats eluted in two distinct peaks (spec. act. = 2.49 and 0.49 nmol/min per mg protein); (c) the elution profile of the enzyme from hepatocyte nodules was broad compared to that from normal liver, surrounding liver or serum; (d) the Km for N-acetyl-D-glucosamine (GlcNAc) was lower in all four independent batches of nodules compared to the Km for GlcNAc from control and surrounding liver; (e) the Km for uridine diphosphogalactose (UDP-Gal) was higher for the enzyme from nodules compared to that from control tissue. These data suggest that the hepatocyte nodule produces several glycoforms of galactosyltransferase the kinetic properties of which differ from those of the enzyme from control liver.