A new heteropolysaccharide, here called P1, was isolated from the fruit clusters of Prunella vulgaris using a hot water extraction method. Chemical and physical analyses indicated that P1 had a spherical conformation with an average molecular weight of 1750 kDa and consisted of arabinose (28.37%), xylose (54.67%), mannose (5.61%), glucose (5.46%), and galactose (5.89%). The main types of P1 linkages were proved to be (1→5)-linked α-L-Ara, (1→)-linked α-L-Ara, (1→3)-linked α-D-xyl, (1→3)-linked β-D-Gal, (1→3,6)-linked β-D-Gal, (1→3,6)-linked α-D-Man and (1→6)-linked α-D-Glc according to the periodate oxidation-Smith degradation and NMR analyses. P1 could significantly enhance the secretion of NO, TNF-α, and IL-6 in murine RAW 264.7 cells, involving the toll-like receptor 2 (TLR2), TLR4 and complement receptor 3 (CR3). Further studies showed that P1 exhibited stable immune activities in the pH range of 4.0-10.0 and below 121 °C. The results suggested that P1 could be used as a potent immunomodulatory agent in functional foods and pharmacological fields.