Optimization of a microplate reader method for the analysis of changes in mitochondrial membrane potential in Chlamydomonas reinhardtii cells using the fluorochrome JC-1

Abstract

The fluorochrome JC-1 is mainly used in mammalian cells to estimate mitochondrial membrane potential (MMP) as a stress marker, with far less data being available for plants or green algae. To address this, we have validated the possibility for changes in MMP to be used as a sensitive stress indicator in the green alga Chlamydomonas reinhardtii. To optimize the method, we analyzed the conditions for applicable MMP determination, including the proper buffer, excitation/emission wavelengths, and solvent (dimethyl sulfoxide, DMSO) influence on the fluorescence signal, and evaluated usefulness of the method for toxicological research. Our results have allowed us to develop a complete protocol for the estimation of MMP changes in C. reinhardtii. Statistical analyses have confirmed the reproducibility, sensitivity, and applicability of this method for toxicological studies.