Abstract: Through establishing an mathematical model, the evaluation of the measurement uncertainty of arsenic and bismuth in gold ore by atomic fluorescence spectrometry (AFS) was investigated. Each partial uncertainty of measurement was analyzed and calculated. The combined standard uncertainty and the expanded uncertainty of arsenic and bismuth were calculated respectively. The uncertainty of As and Bi was mainly composed of the uncertainty of the preparation of samples and uncertainty of the fitting of the standard curve. When the concentration of As in gold ore is 18.398 μg/g, the expanded uncertainty is 0.5553 μg/g (k=2). When the concentration of Bi is 0.3593μg/g, the expanded uncertainty is 0.4411 μg/g (k=2). The expanded uncertainty of Bi is too great to satisfy the requirement of measurement standard. In order to reduce the uncertainty produced by the standard curve fitting, it is necessary to prepare a low concentration of standard solution and increase the mount of samples and reduce the dilution ratio of samples.