Two well-known polysaccharides, (1-->3)-beta-D-glucan and mannan, are major structural components of the fungus cell wall. The G test is a direct method to measure (1-->3)-beta-D-glucan using a (1-->3)-beta-D-glucan-sensitive component, factor G, fractionated from the limulus lysate. The concentration of (1-->3)-beta-D-glucan in culture supernatants of Candida albicans increased to 1,390.0 pg/ml at 24 hours. The concentration of mannan also increased parallel with fungal growth. However, after digestion of supernatants with endo-(1-->3)-beta-D-glucanase, the reactivity to factor G disappeared, although titers of antimannan monoclonal antibody-based latex agglutination were unchanged. Our study demonstrated that cell suspensions of both C. albicans and Cryptoccocus neoformans activated the limulus factor G, and that not only the conidia form but also the filamentous form of Aspergillus fumigatus reacted with factor G. Various Candida spp. (C. paraspilosis, C. glabrata, C. tropicalis, C. krusei), Saccharomyces cerevisiae, Rhodotorula rubra, Trichosporon beigelii, and A. fumigatus released soluble (1-->3)-beta-D-glucan into their culture supernatants, but C. neoformans and Cunninghamella bertholletiae showed only a small reaction to the G test during their culture. Our results indicate that the G test is a good method for serodiagnosis of deep mycosis and also as a screening tool for contamination of medical devices, drugs, and experimental materials with (1 --> 3)-beta-D-glucan.