A new colorimetric method based on paper chip was established for the rapid detection of L-Cysteine（L-Cys） using 5,5'-dithiobis-（2-nitrobenzoic acid）（DTNB） as derivatizing agent.A normal phone was used to record the color intensity.The color intensity was analyzed by Photoshop software and used to calculate the concentration of thiol contained molecules in samples.The optimum coloration conditions for the detection were investigated.Results showed that there was a good linear relationship between color intensity and L-Cys concentration in range of 0.01-0.1 mmol/L with the linear correlation coefficient of 0.999 8 and the detection limit of 0.001 mmol/L.In the range of ±5%,the interference of several amino acids and ions on the determination of L-Cyse was small.The method was applied in the detection of L-Cys content in bovine serum sample with a result of 0.013 mmol/L and a spiked recovery of 99.1%-103%.Results showed that this method has high selectivity and reproducibility.The colorimetric method based on paper based microfluidic chip has several advantages such as simplicity,low reagent consumption,rapidness and repeatability.