In this study, the effects of cold plasma and plasma‐activated medium (PAM) are investigated through different treatment times, different intervals between treatment and analysis, and synthesis of helium gas and helium + 0.5% oxygen. The viability of two cancer cell lines including Hela and MDA‐MB‐231, which are related to cervix and breast cancers, respectively, is investigated using the 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl tetrazolium bromide assay. To study the effect of plasma on normal cells, the corresponding method is used for examination of the normal human fibroblasts. H2O2 showed a higher concentration after direct‐plasma treatment than PAM treatment. Caspase 3, caspase 8, and Bax proteins and the relative Bax/Bcl‐2 ratio are compared in both the direct‐plasma and PAM methods. The results show that the expression level of apoptotic proteins caspase 3 and caspase 8 in direct‐plasma treatment is higher than that in PAM treatment, and the level of relative Bcl‐2/Bax ratio is increased, demonstrating the induction of the apoptosis mechanism.