The effect of the antimicrobial agent TMPAC (3-(trimethoxysilyl)-propyldimethyloctadecyl ammonium chloride) on the cellulase activity on model cellulose substrate was investigated by in situ-null ellipsometry. The cellulases used were extracted from Trichoderma viride and Aspergillus niger, and the model cellulose film was prepared by spin-coating silicon oxide wafers with cellulose solubilized in N-methylmorpholine-N-oxide/dimethyl sulfoxide solution. Upon enzyme addition to the previously equilibrated cellulose film, the initial enzyme adsorption on the substrate was followed by an overall decrease in film mass owing to enzymatic digestion of the cellulose. The loss of cellulose film mass was associated with a non-monotonously behavior of the cellulose film thickness. The activities of the two enzymes were different, a much higher degradation rate being observed for the Trichoderma viride cellulase. The degradation rate with this cellulase decreased significantly when the cellulose film was treated with the antimicrobial agent. The antimicrobial agent did not affect the cellulose degradation catalyzed by the Aspergillus niger cellulase. It was, hence, demonstrated for the first time that, depending on the cellulase type, the antimicrobial agent can inhibit enzymatic activity at the solid-liquid interface.