A polysaccharide (GSP-6B) with a molecular mass of 1.86 × 10⁶ Da was isolated from the fruiting bodies of Ganoderma sinense . Chemical composition analysis, methylation analysis, infrared spectroscopy, and nuclear magnetic resonance spectroscopy were conducted to elucidate its structure. GSP-6B contains a backbone of (1→6)-linked-β-D-glucopyranosyl residues, bearing branches at the O-3 position of every two sugar residues along the backbone. The side chains contain (1→4)-linked-β-D-glucopyranosyl residues, (1→3)-linked-β-D-glucopyranosyl residues, and nonreducing end β-D-glucopyranosyl residues. An in vitro immunomodulating activity assay revealed that GSP-6B could significantly induce the release of IL-1β and TNF-α in human peripheral blood mononuclear cell (PBMC) and showed no toxicity to either PBMC or a human macrophage cell line THP-1. GSP-6B could also activate dendritic cells (DC) by stimulating the secretion of IL-12 and IL-10 from DC.