A new chemically modified electrode (CME) was fabricated, which was based on the immobilization of multi-wall carbon nanotubes fuctionalized with carboxylic group (MWNT-COOH). The results indicated that the CME exhibited efficiently electrocatalytic oxidation for L-cysteine and glutathione with relatively high sensitivity, stability and long-life. Coupled with HPLC, the MWNT-COOH CME was utilized for amperometric detection of the thiols. The peak currents of L-cysteine and glutathione were linear to their concentrations ranging from 3.0×10−7 to 1.0×10−3 mol/L with the calculated detection limit (S/N=3) of 1.2×10−7, 2.2×10−7 mol/L, respectively. The method had been successfully applied to assess the contents of L-cysteine and glutathione in rat striatal microdialysates.