A water-soluble immunoenhancing polysaccharide was isolated from the aqueous extract of fruit bodies of somatic hybrid (Pflo Vv5 FB), obtained through protoplast fusion between Pleurotus florida and Volvariella volvacea strains. On the basis of acid hydrolysis, the polysaccharide was found to contain glucose only. Methylation analysis, periodate oxidation along with (1)H, DEPT-135, and (13)C NMR spectroscopy, including two-dimensional TOCSY, DQF-COSY, NOESY, ROESY, 1H,13C-HMQC, and HMBC experiments showed that the polysaccharide was a (1-->6)-beta-d-glucan, which was not a constituent of any of the parent mushrooms previously reported. This glucan stimulated the macrophages, splenocytes, and thymocytes.