Papain-mediated reduction of MTT is an artifact but not an indicator of cell apoptosis in Saccharomyces cerevisiae

Abstract

Papain is a proteolytic enzyme of the cysteine protease family used for tissue dissociation and cell separation. Papain's nonspecific proteolysis of the plasma membrane enzymes plays a crucial role in the homeostasis by disrupting the intracellular pH of the affected cells which might lead to cell death. When the Saccharomyces cerevisiae cells were treated with different concentrations of papain (1.0, 5.0, 10.0, 20.0, and 30.0 μg/ml), we found no alteration in the trans-plasma membrane electron transport (TPMET) activity and the intracellular pH of the cells, while it significantly decreased the mitochondrial dehydrogenase activity when measured by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay. Additional verification of cell viability by trypan blue assay showed 98%-99% cell viability, contrary to the higher cell death observed with MTT assay. To better understand the decrease in cell viability with MTT assay, we tested the cell-free system that demonstrated a significant decrease of MTT concentration but the trypan blue assay showed more number of viable cells. This study shows that papain interferes with the MTT assay.