The Chinese giant salamander , a member of the family Cryptobranchidae, is the largest extant amphibian species in the world, which is valued for food, medicine, and research on animal evolution and biodiversity because of its unique phylogenetic position and physiological features. Within the last decade, Chinese giant salamander farming has increased rapidly in China. However, with the rapid development of Chinese giant salamander aquaculture, a severe epizootic recently occurred in cultured Chinese giant salamanders in Hubei, Hunan, Sichuan, Shannxi, and Zhejiang Provinces of China, causing tremendous economic losses. The causative pathogen has been identified as the giant salamander iridovirus (GSIV). At present, no effective methods are available for the control of the disease. Immunoprophylaxis is considered the best measure in controlling viral diseases in aquatic animals, and the large-scale cultivation technology of giant salamander cells and GSIV are of significance for the immunoprophylaxis of the disease. In this study, by using Cytodex 3 microcarriers to culture giant salamander muscle cells (GSM) and GSIV at a large scale, the GSM cell morphology, proliferative characterization, and GSIV growth dynamics were investigated. The results showed that the attachment efficacy reached 95% after 10 h of cultivation with intermittent agitation of 30 r/min for 2 min followed by 40 min still culture during the cell attachment period in the Cytodex 3 microcarrier culture system. The optimal culture conditions are as follows:serum concentration 5%, microcarrier concentration 2 g/L, and initial inoculation cell density of 1.2×10⁵ cells/mL. During the cell growth period, the continuous stirring speed was 25 r/min. Under inverted microscopy and electron microscopy, the GSM cells on the Cytodex 3 microcarriers were long, spindle-shaped, and well adhered. After infection with GSIV at a multiplicity of infection of 0.5, the GSM cells on Cytodex 3 microcarriers showed the typical cytopathic effect at 48 h post infection, and the highest virus titer (TCID₅₀/mL) of 10^-8.50±0.20/mL was reached at 72 h post infection. This study established a solid foundation for further investigation on the large-scale technologies of GSIV vaccine production in the future.