As part of a comprehensive effort to evaluate the toxicological potential of calcium sodium metaphosphate fiber (Phosphate Fiber), the in vitro cytotoxicity of the fiber in cultured cells was studied. Two pulmonary-derived cell systems (rat alveolar macrophages, RAM; rat lung epithelial cells, LEC) and an established cell line (Chinese hamster ovary, CHO) were used. Release of lactate dehydrogenase (LDH) was used as an endpoint for cytotoxicity for all three cell types. In addition, inhibition of colony formation was used for CHO cells. The cytotoxicity of Phosphate Fiber was compared to a variety of mineral dusts and fibers including chrysotile asbestos, crocidolite asbestos, two glass fibers, calcium sulfate fiber, titanium dioxide, as well as the nonfibrous raw material, calcium sodium metaphosphate glass. Results with all three cell culture systems demonstrated that the Phosphate Fiber was less cytotoxic than the two asbestos fibers, similar in cytotoxicity to the glass fibers, and more cytotoxic than the calcium sulfate fiber and titanium dioxide. To further investigate the cytotoxicity of the Phosphate Fiber, it was fractionated by sedimentation into small and large fibers. The small Phosphate Fiber was found to be more cytotoxic and the large Phosphate Fiber to be less cytotoxic than the unfractionated Phosphate Fiber. The in vitro data suggest that Phosphate Fiber is less cytotoxic than asbestos, but further determination of safety can only be made after the in vivo data have been obtained.