Label-free mitochondrial dynamics analysis during cell death using organelle-specific phase contrast microscopy (OS-PCM).

Abstract

Mitochondria plays an important role in cell death and undergoes dramatic changes in states of disequilibrium. As mitochondria respond sensitively to cell stress, their dynamics should be studied without affecting cell state. However, current methods rely on labeling cells with fluorescence and introduce additional stress to the cell due to photobleaching and phototoxicity. Here, we propose to use label-free organelle-specific phase contrast microscopy (OS-PCM) to achieve prolonged, specific observation and quantitative analysis of mitochondria dynamics during cell death with minimum perturbation to cells. Using apoptosis and ferroptosis as two examples of cell death, we show quantitatively that large mitochondria tend to increase in size through a combination of swelling and fusion in response to apoptosis, while they decrease in size through fission during ferroptosis. These results provide a new and deeper understanding of mitochondrial dynamics during cell death and demonstrate that OS-PCM is a powerful tool for the gentle, facile, and quantitative study of delicate organelles under stress.