Abstract: To overcome the poor freeze-thaw resistance of conventional protein-stabilized food emulsions, this work prepared Pickering emulsions with excellent freeze-thaw stability using cysteine-rich γ-zein colloidal particles as stabilizers combined with high-pressure microfluidization (HPM). Firstly,γ-zein suspensions with different mass concentrations were fabricated, and their intrinsic physicochemical properties, including particle size, Zeta potential, and protein secondary structure, were characterized to clarify the structural responses of γ-zein particles to concentration changes. Subsequently, emulsions with varying γ-zein contents were fabricated via HPM, and some emulsions were further subjected to thermal treatment. The individual and synergistic effects of γ-zein concentration and heat treatment on the freeze-thaw stability of emulsions were systematically evaluated through three consecutive freeze-thaw cycles (-20 ℃ for 12 h, followed by incubation at 25 ℃ for 2 h). The characterization results showed that increased protein concentration significantly elevated the average particle size and absolute Zeta potential of γ-zein particles, while their overall secondary structure remained relatively stable. Significant differences in the proportion of antiparallel β-sheets were observed only at the γ-zein mass concentration of 30 mg/mL. HPM treatment remarkably improved the freeze-thaw tolerance of the fabricated emulsions. No oil separation was detected in any emulsion samples after three freeze-thaw cycles, and only the emulsion stabilized by 10 mg/mL γ-zein exhibited slight creaming behavior. In addition, thermal treatment exerted a negligible influence on the freeze-thaw stability of HPM-treated emulsions. Further mechanistic analysis revealed that HPM treatment effectively strengthened the structure tolerance of emulsion systems to structural damage triggered by ice crystal formation and growth. The synergistic interaction between high γ-zein concentration and HPM treatment promoted the formation of compact oil-water interfacial films and continuous three-dimensional particulate networks in the aqueous phase, which was responsible for the improved freeze-thaw resistance of emulsions. Although thermal treatment slightly weakened the adsorption of γ-zein particles at the oil-water interface and within the bulk network structure, the resultant emulsions still maintained satisfactory freeze-thaw stability, which meets the requirements of practical food applications.