Collapse of the proteostasis can lead to accumulation and aggregation of unfolded proteins, which has been found to associate with a number of disease conditions including neurodegenerative diseases, diabetes and inflammation. Here we report a maleimide-functionalized tetraphenylethene (TPE)-derivatized fluorescent probe, which shows fluorescence turn-on property upon reacting with unfolded proteins in vitro and in live cells under proteostatic stress conditions. The level of unfolded proteins can be measured by using flow cytometry and visualized with confocal microscopy. Furthermore, it can be used to enrich unfolded proteins in cells. Proteomics approach identified and quantified unfolded proteins in different stress conditions.