Latex allergies are caused by latex protein and the chemicals such as accelerators added during the manufacturing of its products. Cytotoxicity and hypersensitivity type I and type IV reactions are the common clinical manifestations of natural rubber latex (NRL). The study is to evaluate the cytotoxic effect of medical gloves prepared by diisopropyl xanthogen polysulfide (DIXP) and zinc diisononyl dithiocarbamate (ZDNC), which are claimed to be safer than conventional accelerators towards HaCat cells. Gloves prepared with varying concentrations of DIXP and ZDNC were subjected to cytotoxic tests, specifically agar overlay assay, filter diffusion test and MTT test, which HaCat Cell lines were used as the in vitro model. The tests showed that all the gloves prepared with DIXP (1.0, 1.2 and 1.5 phr) shows mild cytotoxicity while the gloves prepared with 1.0 phr of ZDNC has no cytotoxic effect in the agar overlay and filter diffusion tests. However, gloves prepared with 1.2 and 1.5 phr of ZDNC showed mild cytotoxicity in both tests. As for the MTT test, both DIXP and ZDNC gloves regardless of the concentration, showed a steady cell viability above 80% before decreasing to less than 20% at higher concentrations of 50% and 100% of sample concentration prepared by two-fold serial dilution. Taken together, the results suggest that ZDNC is a more viable and safer alternative for replacing conventional accelerators. Further testing is necessary to confirm the cytotoxicity of ZDNC particularly at a concentration not exceeding 1.0 phr.