Gold nanoclusters (AuNCs) were synthesized by a macromolecules template using bovine serum albumin (BSA) as stabilizer which can emit red photoluminescence under illumination of ultraviolet light. The fluorescence intensity of AuNCs enhanced through decreasing the surface defects of AuNCs modified with cysteine, herein we present a novel fluorometry for determination of trace cysteine. This method with a wider linear range from 2.0 to 800 nmol mL(-1), higher sensitivity (detection limit was 1.2 nmol mL(-1)) and better selectivity has been utilized to determine cysteine content in real samples, and the results were in a good agreement with those determined by electrochemical biosensor. At the same time, the structures of AuNCs and AuNCs-cysteine were characterized by Fourier-transform infrared spectroscopy (FTIR) and high resolution transmission electron microscopy (HRTEM) and the mechanism of the proposed assay for the detection of cysteine has been discussed.