Label-free fluorometric assay for cytochrome c in apoptotic cells based on near infrared Ag₂S quantum dots

As an important biomarker, cytochrome c (Cyt c) plays a crucial role in mitochondrial electron transport chain and cell apoptosis. Herein, a label-free near-infrared (NIR) Ag₂S quantum dots (QDs)-based fluorescent strategy was constructed for the sensitive and selective detection of Cyt c. In this system, Cyt c was hydrolyzed by trypsin, and the resulting heme-peptide fragment exhibited peroxidase-like activity for catalytic decomposition of H₂O₂ into hydroxyl radical (·OH). The presence of caffeic acid in this system resulted into the formation of caffeic acid-quinone due to the strong oxidizing ability of ·OH. The production of caffeic acid-quinone led to the fluorescence quenching of Ag₂S QDs through electron transfer mechanism. Based on the cascade reaction, we successfully developed a label-free approach to detect Cyt c using Ag₂S QDs as nanoprobes. Under the optimized conditions, the fluorescence intensity of Ag₂S QDs was linearly relative to the concentration of Cyt c over the range from 2.0 to 150 nM with a detection limit of 1.7 nM. In addition, this strategy was successfully applied for quantitative detection of Cyt c in cell lysates of H₂O₂ or etoposide (anticancer drug)-induced apoptotic cells, providing great potential application for cell-based oxidation pressure determination and screening of anticancer drugs.