The change pattern of blood cholinesterase activity among organophosphorus pesticides workers varies with acute or chronic exposure manner. In this paper, the significance of blood cholinesterase activity assay, serving as a surrogate effect biomarker to reflect the neurotoxicity in target cholinergic nerve tissues due to acute exposure to organophosphorus pesticides, and its application values in occupational medicine clinical practice were introduced; and multiple time-related influencing factors relevant to this assay such as duration and time point of exposure, monitoring time period, stage of disease course, and phase of enzyme activity regeneration process were also analyzed. Meanwhile, the characteristics of enzyme activity and derived issues in occurrence of cholinergic neurotoxicity tolerance after long-term low-level exposure to organophosphorus pesticides and the considerations of this assay for occupational health surveillance were described.