Vibrio spp. is one of the main pathogens to cause high levels of mortality in the culture industry of mud crabs, Scylla paramamosain. Although considerable efforts have been devoted to understanding the immune response of S. paramamosain against pathogens, both immune‐response proteins and systematic responses to foreign agent invasions have yet to be investigated in detail. Hence, in this study, we challenged S. paramamosain with bacterial lipopolysaccharides (LPS) by injection. We examined a number of immune‐related indices in the crabs at 0, 2, 6, 12, 24, 36, 48, and 72 h postinjection, that is, the total hemocyte counts in hemolymph; the serum activities of phenoloxidase, superoxide dismutase, lysozyme, acid phosphatase, and alkaline phosphatase; and the mRNA levels of immune‐related genes encoding prophenoloxidase, serine protease inhibitors, antimicrobial peptide scygonadin, crustin, anti‐lipopolysaccharide factor, and catalase in hemocytes. In addition, the immune‐response proteins were identified in hemocytes at 24 h postinjection using two‐dimensional polyacrylamide gel electrophoresis. The results showed that the immune‐related enzymes and genes examined were induced in a time‐dependent manner post‐LPS challenge. Moreover, 18 altered proteins were present in the hemocytes and the main proteins to be altered were highly homologous to hemocyanin and protein kinase. Ultimately, this study provides useful information for understanding the immune response of S. paramamosain to bacterial infection.