Naturally, refractory gold is enclosed by sulfide minerals such as pyrite and arsenopyrite and a higher content of organic carbon, which results in difficulty in the recovery of gold. In this study, the flotation gold concentrate before and after pressure oxidation (POX) was analyzed. The analysis of the Eh-pH diagram revealed that pyrite/arsenopyrite would be dissolved in higher oxidation potential. Results of SEM-EDS showed that the surface of minerals became very rough and structure of some particles was destroyed after POX. The XRD measurement showed that pyrite was oxidized to jarosite, and gangue minerals such as quartz and organic carbon were hardly changed. The process of thiosulfate leaching of gold was designed and optimized according to Miscellaneous implemented by Response Surface Methodology (RSM). Based on the results of ANOVA, leaching model was significant. In leaching, [S2O32-] played an important role and the interactions between [S2O32-] and [Cu2+] were significant. The effect of [HA] was relatively weak. Optimum process parameters of the initial reagent concentration were 505.00 mM for [S2O32-], 34.77 mM for [Cu2+], and 0.50 mM for [HA]. Under these conditions, the Au extraction value was about 86.21%, which was significantly higher than that obtained with leaching directly.