The polysaccharide from from roots of Radix Tetrastigma were extracted by water bath, and the yield of polysaccharide was determined by response surface methodology. Based on the single factor test, The optimum conditions were as followed: temperature 96 ℃, ratio of water to liquid 34∶1 （mL/g）, extraction time 2 h. Under the conditions, the yield of polysaccharide was (21.23±0.77)%. After deprotein with sevage method, fractionated and purified by DEAE-Cellulose 52 anion exchange column and Sepharose CL-4B gel fltration column, successively . The purifed polysaccharide was designated as RTP-3-1 and its components, molecular weight, primary structure and domains were identified by high performance gel permeation chromatography (HPGPC), fourier transform infrared spectroscopy and hydrogen nuclear magnetic resonance spectroscopy (HNMR). The results showed that RTP-3-1 was high purity polysaccharides with a molecular weight of 1 244.2 kDa，and had the characteristic absorption peak of acidic polysaccharide. ¹HNMR showed RTP-3-1 was an α-type polysaccharide.