In this study, a novel heteropolysaccharide named SP90-1 with immunostimulatory and antitumor activity was purified and characterized from Spirulina platensis. SP90-1 has a molecular weight of 63.92 kDa and mainly consists of rhamnose (Rha), glucose (Glc), galactose (Gal) and glucuronic acid (GlcA), followed by the minor components Fuc and Xyl. The backbone of SP90-1 was determined to be →2)-α-d-Rhap-(1 → 2,3)-α-d-Rhap-(1 → 4)-β-d-Glcp-(1 → [3)-β-d-Rhap-(1→], with branches at the O-3 of Rha, consisting of the side chains 4-Galp and 4-GlcpA. SP90-1 was found to significantly enhance phagocytic capacity, promote the secretion of nitric oxide (NO), interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) in RAW264.7 cells, and remarkably inhibit the growth of A549 lung cancer cells. These findings demonstrate that SP90-1 could potentially be further explored for immunomodulatory biomedical applications.