These studies were carried out to investigate the properties of H-Y antigen purified by immunoaffinity chromatography using monoclonal H-Y antibody. Immunoaffinity column was prepared by the coupling of monoclonal antibody to the Aminolink Coupling Gel. Murine testis supernatant was applied onto the column and eluted by O.lM glycine-HCl buffer and 31${mu}g$ of H-Y Ag was eluted from one testis. Purified H-Y Ag strongly reacted with Con A and lentil from 6 different kinds of lectins tested, which may indicate that sugar moiety of H-Y Ag is composed of glucose, mannose and their derivatives. Con A-sepharose affinity column was used to purified H-Y Ag based on that H-Y Ag is glycoprotein. The fraction eluted by 0.2M Me-${alpha}$-D-mannoside from the column loaded with murine testis supernatant was identified to be H-Y Ag by dot blot test. Molecular weight of the purified H-Y Ag was estimated by Sepharose G-75 gel filtration and SDS-PAGE, and showing that it was about 67,000 dalton. In fluorescence test, the ratio of XY embryos and XX embryos was 1:1.