The chemical cross-linking of soluble (1→3)-β-D-glucans having molecular weights of 21000 (CL 3h) and 6400 (CL 6h), and laminarin (CL LAMI), which showed negligible biological activity, by epichlorohydrin provided rigid particles. These particles showed no gel-to-sol transition upon the addition of sodium hydroxide. We compared the effects of chemical cross-linking on the biological activities of glucans. The alternative complement pathway was not activated by any of the cross-linked glucans. Glucose consumption, lysosomal enzyme release, and interleukin-1 production by mouse resident peritoneal macrophages incubated in vitro were strongly induced by CL 3h, CL 6h and CL LAMI. However, cross-linked dextran, Sephadex, did not exhibit any of these biological activities. These results suggested that chemical cross-linking of (1→3)-β-D-glucans enhances macrophage activities without opsonization by complement components.