The present study compares three specific toxicity parameters for cytotoxicity testing of chemically different dental materials. Two glass ionomer cements, a zinc phosphate cement, and a composite material were used to evaluate the sensitivity of three assays: two viability assays, the MTT assay and the quantifiable neutral red assay, and a proliferation assay based on the determination of the total protein content of a cell culture. The colorimetric assays were carried out using transformed mouse fibroblasts (L-929 cells) and fibroblasts derived from biopsies of normal human gingiva. In most cases, all colorimetric assays detected much weaker cytotoxic responses, if any, in gingival fibroblasts than in L-929 cells. The viability assays indicated cytotoxicity of the extracts to two glass ionomer cements in L-929 cells when the materials were set at 0% relatively humidity for 24 h. The severe cytotoxicity of the zinc-phosphate cement in both viability assays was less influenced by the setting conditions. The cytotoxicity of the composite material was most pronounced in the neutral red assay. In general, both the MTT assay and the neutral red assay were more sensitive than the colorimetric proliferation assay. These assays can be performed very effectively; only few cells are needed for rapid, reliable and inexpensive screening purposes of a large number of samples in a short time. Automated processing with a microplate reader after non-radioactive labeling of the cells and subsequent automated analyses of original data, with no need for sophisticated and expensive equipment, are additional advantages of the systems.