2001 Volume 36 Issue 2 Pages 96-98
A simple assay to determine the proliferation of larval and juvehile fish splenocytes using Alamar Blue (AB) was studied. Splenocytes were separated by Percoll gradient from Japanese flounderParalichthys olivaceus (1-year-old), then the proliferation of splenocytes stimulated by mitogen (concanavalin A : ConA, pokeweed mitogen : PWM, or lipopolysaccharide : LPS) was detected using AB. The relationship between number of splenocytes and specific absorbance exhibited a positive significant correlation. The optimum condition of this assay was 5×105 cells/well of splenocytes with mitogens (ConA 100μg/mL, PWM 10μg/mL or LPS 1μg/mL) for 72 h. Proliferation of each splenocyte from juveniles of Japanese flounder and larvae of Japanese parrotfish Ophegnathus fasciatus and tiger puffer Takifugu rubripes was detectable by this assay.