Abstract: Aim to evaluate the uncertain factors which may influence the accuracy of detection of pesticide residues in fresh tea leaves, a case study of dichlorvos, dimethoate, malathion and fenitrothion residues in fresh tea leaves was carried out using gas chromatography with external standard method. According to the relevant standard and protocol, a mathematical model has been studied. The uncertainties created by the entire experimental process were evaluated systematically. Three A-type evaluation components, including reproducibility of determination, fitness of standard curve and recovery rate, and four B-type evaluation components, including preparation of standard solution, sample weighing and pretreatment, and the measurement of instrument were investigated. The results revealed that the expanded uncertainty of measured values of dichlorvos, dimethoate, malathion and fenitrothion in the spiked tea leaves were 0.021, 0.073, 0.036 and 0.023 mg/kg (k=2, confidence interval P=95%), while the theoretical content were 0.096, 0.39, 0.19 and 0.19 mg/kg, respectively. Preparation of standard solution and fitness of standard curve contributed to the major effect on uncertainty, which should be paid more attention for future determination. Our study provides a reference for the accuracy assessment of method in detection of four organophosphorus pesticide residues in fresh tea leaves.