In order to investigate the differentiation change of tissue proteins in fish after infection with Vibrio mimicus, Carassius auratus were artificially infected by V. mimicus 04-14 isolates. Differential proteome analysis in spleen and intestinal mucosa tissues before and after infection was performed by using two-dimensional polyacrylamide gel electrophoresis (2-DE) combined with matrix assisted laser desorption/ionization time-of-flight mass spectrometry techniques (MALDI-TOF-MS). The results showed that there were 11 and 12 significantly differentially expressed proteins in spleen and intestinal mucosa tissues after infection, respectively. A total of 19 significantly differentially expressed proteins were identified by MALDI-TOF-MS, with 10 up-regulated and 9 down-regulated proteins. Amylase, apolipoprotein A-I, α-actin and peroxiredoxin 2 were the common significantly differentially proteins both in spleen and intestinal mucosa tissues. Gene Ontology (GO) annotation indicated that significant differentially expressed proteins were located in extracellular region, cytoplasm, mitochondrion and reticulum, respectively. The molecular functions of these proteins included catalytic activity, binding, transport, immune, antioxidant, cytoskeleton and membrane skeleton formation, involved in eight biological processes of substance and energy metabolism, cell membrane transport, complement activation, response to stimulus, cytoskeleton formation, protein folding, redox and iron ion transport. These results might provide a basis for further research of V. mimicus pathogenic mechanism and bacteria- host interactions.