Dentinogenically active rat incisor odontoblasts were dissected out from animals fed a low calcium, vitamin D free diet (R 25). Protein synthesis by these cells was studied by means of short-time incubation in an in vitro system in the presence of the radioactive labeled precursors L-leucine, L-fucose and L-proline. A significantly increased leucine and proline incorporation into the protein synthesized was noted in the cells from rats fed the deficient R 25 diet compared with odontoblasts from rats fed an adequate control diet (R 47). No difference between the two groups was found when fucose was utilized as a precursor. The well known increase in predentin width when feeding a rachitogenic diet may thus be explained by an increase in organic matrix synthesis in addition to the possible negative direct effects of lowered serum Ca content. Prior to this study, the behavior of proline as a precursor in the in vitro system was studied. The possibility of separating leucine-labeled proteins synthesized in the in vitro system by means of SDS-polyacrylamide gel electrophoresis was also shown.