Objectives: The aim of this study was to explore the involved pathophysiological processes and develop biomarkers of trichloroethylene-induced hypersensitivity dermatitis (THD). Methods: We examined the impact of THD on the serum proteome in 8 male patients by comparing the serum samples between acute and healed stages. Sample pooling and immunodepletion were applied for sample preparation. Two-dimensional gel electrophoresis coupled with matrix-assisted laser-desorption ionization time-of-flight mass spectrometry (MALDI-TOF-TOF/MS) was utilized to identify and quantitate differentially expressed proteins. Changes in selected proteins were further confirmed by an ELISA assay. Results: A total of 41 spots were quantitated with significant alteration ( p <0.05; fold-change≥± 3.0) in the serum between the acute and healed stages. Of these proteins, 26 proteins were identified by MALDI-TOF-TOF/MS. The identified proteins could be categorized into diverse functional classes, e.g., immunity and defense response, vitamin and lipid transport, fatty acid biosynthesis, actin binding, proteolysis and glycolysis. The ELISA assay confirmed the relative upregulation of calprotectin (S100A8/A9) and downregulation of retinol binding protein (RBP4) in the serum of the acute stage. The alteration of calprotectin and RBP4 was found to be specific to THD rather than trichloroethylene exposure. Conclusions: The pathophysiological processes underlying THD may involve elevated inflammatory responses and oxidative stress, inhibition of vitamin transport, depression of fatty acid biosynthesis, loss of extracellular actin scavenger, increase in oxygen transport, dysfunction in lipid transport, proteolysis and glycolysis. The combination of higher calprotectin and lower RBP4 levels in the serum could be used as potential biomarkers of THD.