Primary cultured cells derived from human pulp tissue were used for examining the cytotoxicity of nine commercial filling materials used in restoration and root canal procedures. Cell growth was evaluated by photo pattern analysis, cell nuclei counting, and neutral red uptake. L-strain cells were also used for comparison. Primary cultured cells from either the coronary or radicular part of the pulp tissue were less sensitive to the tested materials, than were L-cells. As for sensitivities over time following mixing of the materials, the cytotoxicity of the three restorative materials was immediately moderate but disappeared 24 hours after mixing. The other six root canal filling materials yielded similar moderate to high cytotoxic levels immediately after mixing and the effects continued. The order of cytotoxicity was: Diaket and Calvital (highest); Pulp canal sealer, AH-26, and Canals (high); and then Kloroperka and three restorative materials (moderate). The differences in cellular reactions between primary and strain cells was considered to be due to cell origin, i.e. diploid or heteroploid. The importance of using primary cultured cells from designated target tissues when evaluating various materials in vitro was discussed.