A method was developed for the purification and detection of selenoprotein-P in human plasma.The sample was gradient eluted by two successive steps of affinity chromatography,i.e.,Heparin-Sepharose and Ni-Sepharose chromatography.The quality of the purified selenoprotein-P was detected by HG-AFS.The optimum gradient elution conditions were determined and the selenoprotein-P with a certain purity was obtained by SDS-PAGE.The recovery of the two steps of affinity chromatography reached up to 43.2%.The correlation coefficient of HG-AFS was 0.999 1，and the detection limit was 0.09 μg/L.The RSDs were 0.12% for within-day and 0.27% for inter-day.The recoveries were in the range of 95% - 104%.The proposed method was simple and sensitive,and the results obtained were accurate.