To research the relationship between structural characterization and immunomodulatory activities of a polysaccharide fraction， a polysaccharide fraction was isolated and purified from Sparassis latifolia polysaccharides and its structure and immunomodulatory activities was investigated. The polysaccharide fraction SlP was isolated and purified from Sparassis latifolia polysaccharides by HZ-830 macroporous resin， DEAE-52， and Sephadex-G100 chromatography. The structure of SlP was investigated using high performance gel permeation chromatography， infrared spectroscopy， ion chromatography， nuclear magnetic resonance， atomic force microscopy， scanning electron microscopy and Congo red. The MTT method was used to measure the proliferation activity of macrophages RAW264.7 neutrophic red staining was used to determine the phagocytosis ability of SlP. Cell supernatants were collected and the production of NO in the supernatant were detect. Cells were collected and total RNA was extracted， then the expression levels of TNF-α， IL-1β， IL-6， IL-3， IL-10 and IFN-β mRNA in macrophages were determined using quantitative RT- PCR. SlP exhibited a triple helical structure with β-D-glucan and its average molecular weight was 1.04 × 104 u. The surface microstructure of SlP exhibited different shapes and sizes of fragments. The molecular morphology revealed that SlP molecules were compact and asymmetric in shape with branching and entangled chains. SlP could promote RAW 264.7 macrophage proliferation at the 1.95-31.25 μg/mL concentration range； in particular， proliferation reached a peak at 7.81 μg/mL SlP. At the 1.95-7.81 μg/mL concentration range， SlP significantly improved the phagocytic capacities of RAW264.7 macrophages， increased the production of NO， and increased tumor necrosis factor （TNF）-α， interleukin （IL）-1β， IL-6， IL-3， IL-10， and interferon （IFN）-β mRNA levels. Thus， SlP could be used as good functional food raw materials in food industries.