ZnS nanoparticles have been prepared and modified with sodium thioglycolate. The functionalized nanoparticles are water-soluble. They were used as fluorescence probes in the determination of proteins, which was proved to be a simple, rapid and specific method. In comparison with single organic fluorophores, these nanoparticle probes are brighter, more stable against photobleaching, and do not suffer from blinking. Under optimum conditions, linear relationships were found between the enhanced intensity of fluorescence at 441 nm and the concentration of protein in the range 0.1–4.0 μg mL⁻¹ for human serum albumin (HSA), 0.2–3.0 μg mL⁻¹ for bovine serum albumin (BSA) and 0.1–4.5 μg mL⁻¹ for γ-globulin (γ-G). The limits of detection were 0.015 μg mL⁻¹ for HSA, 0.024 μg mL⁻¹ and 0.017 μg mL⁻¹ for BSA and γ-G, respectively. The method has been applied to the analysis of human serum samples collected from the hospital and the results were in good agreement with those reported by a hospital, indicating that the method presented here is not only sensitive and simple, but also reliable and suitable for practical application.