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[Display omitted] • Two generations of sphalerite were identified in the Ashele Cu-Zn ores. • The ore-fluid evolution at Ashele is recorded by sphalerite geochemistry. • Sulfur isotopes show a magma and TSR-derived sulfur source. • Sulfide geochemistry has the potential to guide mineral exploration. The Ashele Cu-Zn deposit is one of the largest and most typical VMS-type Cu-Zn deposits in the Chinese Altai Mountains, Central Asian Orogenic Belt. The ore-forming processes of the Ashele deposit remains elusive, with particular uncertainties surrounding the physicochemical conditions underpinning their formation. In this study, major and trace element concentrations of the two types of sphalerite and sulfide sulfur isotopic compositions have been analyzed to constrain the mineralization conditions and ore genesis. Two stages of sphalerite have been identified, i.e., euhedral sphalerite of the early Cu mineralization stage (Sp1) and anhedral sphalerite of the late Zn mineralization stage (Sp2). Results show that Sp1 has relatively higher contents of In, Cu, Sn, and Hg, but lower levels of Cd, Ga, Ge, As, and Ag compared to that of Sp2. Utilizing the sphalerite GGIMFis geothermometer, mineralization temperatures for Sp1 and Sp2 were estimated at 225-296°C (average 258°C) and 166-264°C (average 210°C), respectively. Additionally, the Ashele sphalerite geochemistry suggests a relatively high sulfur fugacity during ore formation (l g f S 2 = −8.99 to −6.55 for Sp1 and −13.58 to −6.83 for Sp2). The findings suggest an evolutionary trend of ore-forming fluids, characterized by a decrease in both temperature and sulfur fugacity from the early Cu mineralization stage to the later Zn mineralization stage. In situ δ34S values of sulfides (sphalerite, chalcopyrite, and pyrite) range from 0.84 ‰ to 6.07 ‰, indicating a sulfur origin from the thermochemical reduction of seawater sulfate and leaching of the underlying volcanic rocks, with a minor contribution from direct gas release from shallow magma chambers. Overall, this work unravels the physicochemical transformation between the two identified mineralization stages, thereby enhancing the understanding of the ore-forming process of the Ashele deposit. [ABSTRACT FROM AUTHOR]