Filamentous fungi are widely used to produce cellulase, but how the fermentation conditions affect their production is not well known. In this study, cellulase production by Trichoderma viride in submerged fermentations with free cells in a stirred-tank reactor (STR) and immobilized cells in a rotating fibrous-bed bioreactor (RFBB) were investigated. Compared to free-cell fermentation, immobilized-cell fermentation gave 35.5% higher FPase activity and 69.7% higher saccharification yield of sugarcane bagasse (SCB). The secretory proteins in the fermentation broths were analyzed with two-dimensional gel electrophoresis (2-DE) and MALDI-TOF-TOF mass spectrometry, which identified 24 protein spots with differential expression levels. Among them, cellobiohydrolase CBH II and endoglucanase EG II were highly expressed and secreted in the immobilized-cell fermentation, while the free-cell fermentation produced more CBH I and EG IV. These results showed that immobilized-cell fermentation with T. viride in the RFBB was advantageous for cellulase production. (C) 2013 Elsevier Ltd. All rights reserved.