Abstract: Large yellow croaker (Larimichthys crocea) is an important economic species in Fujian. To develop virological, toxicological and cell engineering techniques, as well as resources for advanced researches on the fish, this study had developed a new cell line derived from the fish kidneys(YCK). The obtained YCK has been successfully conserved in the M-199 medium supplemented with 10% fetal bovine serum at 27℃ over 400 days with a medium-change every 72 h, and satisfactorily sub-cultured more than 70 times during the interim. A parallel test of commonly applied culture media was conducted to get the conclusion that M-199 being the most suitable medium for YCK. This newly established cell line consisted predominantly of epithelial-like cells, and was proven to be diploid with a modal chromosome number of 2n=48.