<p>Mass mortality of mandarin fish (<i>Siniperca chuatsi</i>) due to infectious spleen and kidney necrosis virus (ISKNV) infection occurs frequently. Since there are no effective drug treatments available, prevention relies heavily on detection. Effective and rapid on-site detection methods are needed for early diagnosis of ISKNV. In this study, a rapid and simple colloidal gold test strip method, specific for the antibody against major capsid protein (MCP), was developed and systematically evaluated for the detection of ISKNV. The limit of detection of the test strip is a 1:100 dilution of a positive standard serum and the antibody level in the fish could be estimated from the depth of color of the test line. The strips were tested against serum samples of cyprinid herpesvirus-2 (CyHV-2), grass carp reovirus (GCRV), largemouth bass ranavirus (LMBV), large yellow croaker iridovirus (LYCIV), and spring viremia of carp virus (SVCV), yielding no cross-reactivity. In addition, 10 mandarin fish artificially infected with ISKNV were tested using the current industry standard PCR method (SC/T 7211-2011) on their splenorenal tissues. The results from the test strips showed a high degree of concordance with PCR testing, achieving a Kappa value of 0.737. All the results indicated that the colloidal gold test strips prepared in this study could be used as a simple, rapid, and highly sensitive and specific method for ISKNV diagnosis.</p>