Based on the chemiluminescence reaction of KMnO4-HNO3-2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside sensitized by HCHO, an analytical method for 2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside by RP-HPLC with chemiluminescence detector was developed. The chromatographic conditions were optimized. AN ODS hypersil column(25 mm x 4.0 mm, 125 x 4.0 mm i.d. 5 microm) was used with acetonitrile--0.3% H3PO4 [25:75, (phi)] as mobile phase, and the flow rate was 0.8 mL x min(-1) at the column temperature of 40 degrees C. The linear range of calibration curve was from 15.75 to 136.5 microg x mL(-1), the detection limit was 11.83 microg x mL(-1) with a concentration of 21 microg x mL(-1) of 2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside, and the recovery was 102%-108%. The relative standard deviation was 3.45% (Cs = 21.0 microg x mL(-1), n = 3). The method has been applied to the determination of 2,3,5,4'-tetrahydroxystilbene-2-O-beta-D-glucoside in a few Chinese medicines, and the results were satisfactory. The method was simple, rapid and highly selective.