Abstract: To investigate the epidemiology and etiology of eel viral diseases, isolate, cultivate and identify the eel virus, a new cell line (EEK) was established by tissue explant method from European eel Anguilla anguilla. EEK cells were fibroblast-like cells and were maintained and subcultured 38 generations over a 12-months period. The optimization of medium, serum concentration and temperature was conducted. EEK cells can grow and proliferate normally in DMEM/F12 and L15, but can not proliferate in MEM. The growth rate enhanced with the increased Fetal Bovine Serum (FBS) in the range of 5%—15%, and it decreased with more than 20% FBS and less than 5% FBS. The cells grew well in a temperature range of 22—27℃, but the growth rate reduced at the temperature below 17℃ or above 32℃. The virus sensitivity test revealed that the cell line was susceptible to the infection of Anguillid herpesvirusv (AnHV) and has obvious cytopathic effect. The establishment of a kidney cell line from european eels has increased the variety of fish cell lines, providing important experimental material for the diagnosis of eel viral diseases, the study of viral etiology and the development of virus vaccine.