Syneresis and low viscosity are two major quality defects found in yogurt. The addition of food stabilizer is necessary to overcome the issue. Oyster mushroom (Pleurotus ostreatus) could be utilized as a source of natural food stabilizer because it contains β-glucan. The objective of this study was to evaluate the effect of Pleurotus ostreatus aqueous extract (POAE) on physicochemical properties, protein profile and total lactic acid bacteria of yogurt fortified with Lactobacillus acidophilus. Yogurt was processed with the addition of POAE at 1%, 2%, and 3% (v/v) before fermentation and compared with control. Yogurt added with 3% POAE had the highest viscosity (P<0.05), protein content (P<0.05), and total acid content (P<0.05), but had the lowest syneresis (P<0.05) among others. The addition of POAE up to 3% (v/v), however, did not affect reducing sugar content and total lactic acid bacteria in yogurt. A 12.49 kDa protein was present in yogurt added with 3% POAE. Therefore, POAE could be used as natural stabilizer for yogurt fortified with Lactobacillus acidophilus with recommended addition level of 3% (v/v).