Orogenic gold deposits are the most important gold resource in the world. It is controversial whether their ore-forming fluids were sourced from metamorphic dehydration of the crust or the mantle. The Tokuzbay gold deposit is a typical orogenic-type gold deposit in the Central Asian Orogenic Belt, providing a case for discussing the source of ore-forming fluids. The auriferous quartz veins are fault-controlled and occur mainly in metamorphic sediments of Devonian Altai Formation and minor in diorite dikes, forming three ore types, i.e., quartz vein, altered metamorphic sediments and altered diorite dikes. We studied the Sr–Nd–Pb isotope characteristics of the ore sulfides, and magmatic and metamorphic rocks from the Tokuzbay gold deposit, and thereby discuss the source of the ore-forming fluids. The 87Sr/86Sr ratios of meta-sediment hosted ores scatter between 0.709474 and 0.715238, and ISr (292 Ma) values range from 0.706570 to 0.714045. The diorite dikes hosted ores show more restricted 87Sr/86Sr ratios ranging from 0.706963 to 0.711519, and ISr (292 Ma) values ranging from 0.706777 to 0.710612. The wall rocks of the Altai Formation have ISr (292 Ma) values of 0.711778 and 0.714409, which are higher than ore sulfides. This indicates that the ISr (292 Ma) values of initial ore-forming fluids that interacted with the Altai Formation are no higher than 0.706777, which is the lowest ISr (292 Ma) value for disseminated ores. The calculated (143Nd/144Nd)i ratios and εNd (292 Ma) values for meta-sediment hosted ores range from 0.511882 to 0.512464, and −7.4 to 4.0, respectively. The calculated (143Nd/144Nd)i ratios and εNd (292 Ma) values for diorite dike hosted ores vary from 0.512293 to 0.512562, and from 0.6 to 5.9, respectively. The meta-sediments of Altai Formation show εNd (292 Ma) values ranging from −6.8 to −4.4 with an average of −5.8, which are lower than those of ore sulfides. This indicates that the εNd (292 Ma) values for the fluids that interacted with the wall rocks cannot be lower than 4.5, which is the average value for disseminated ores. The calculated (208Pb/204Pb)i, (207Pb/204Pb)i, and (206Pb/204Pb)i values of ore sulfides are 37.796–38.989, 15.512–15.707, 17.988–18.100, respectively. The wall rocks of the Altai Formation show relatively higher (206Pb/204Pb)i ratios, indicating that the ore-forming fluid that interacted with the Altai Formation are not higher than 17.988, which is the lowest values of sulfides from disseminated ores. Based on the Sr–Nd–Pb isotope data, we suggest the possible ore-forming fluids should have lower ISr (292 Ma) values, higher εNd (292 Ma) values, and lower (206Pb/204Pb)i values compared to the those of ore sulfides and were possibly sourced from a source compositionally similar to the diorite dikes in addition to wall rocks of Altai Formation. Combining with the regional metamorphism and tectonic evolution, we suggest that the ore fluids have originated from the metamorphic dehydration of sediments of Altai Formation in the South Altai during the collision between Altai Orogen and Junggar Plate.