Cold atmospheric plasma has emerged as a novel oncotherapeutic approach that has undergone a fast growth in the past few years. However, there is no quantitative assessment of the factors influencing its anti‐tumor efficacy and the mechanism driving its activity remains mysterious. Through the use of 18 orthogonal experiments followed by linear model construction, we identified four deterministic parameters, that is, “treatment time,” “liquid surface area,” “thickness of medium,” and “number of cells,” which play deterministic roles on the anti‐tumor efficacy of plasma. We propose “Reactive Species Diffusion Model” and “Signal Transduction Model” to explain the mechanics determining the activity of plasma treated medium and its cell‐death induction efficacy. We, in addition, proposed the use of deterministic parameters in defining plasma dose in the liquid form to more objectively reflect its multi‐modality nature. Our study contributes in elucidating plasma activity and efficacy in a quantitative way, which guides other plasma related investigations and accelerates plasma clinical applications as a type of precision oncotherapy.