Mitochondrial quality control is paramount for cellular development, with mitochondrial electron flow (Mito-EF) playing a central role in maintaining mitochondrial homeostasis. However, unlike visible protein entities, which can be monitored through chemical biotechnology, regulating mitochondrial quality control by invisible entities such as Mito-EF has remained elusive. Here, a Mito-EF tracker (Mito-EFT) with a four-pronged probe design is presented to elucidate the dynamic mechanisms of Mito-EF's involvement in mitochondrial quality control. Heightened aggregation of Mito-EF in fiber-like healthy mitochondria compared to round-like damaged mitochondria is demonstrated, revealed Mito-EF aggregation correlated with mitochondrial morphological remodeling, particularly in regions undergoing mitochondrial fission and fusion, and show the Mito-EF signal associated with mitochondrial cristae maintained by Dynamin-Related Protein 1 (DRP1). This underscores the importance of considering Mito-EF in assessing mitochondrial quality control parameters. A novel drug screening evaluation parameter, Mito-EF is also introduced to screen and discover mitochondrial-targeted therapeutic modulators. This tracker provides new avenues for investigating the role of Mito-EF in maintaining mitochondrial homeostasis and quality control, offering a potent tool for assessing mitochondrial quality and drug screening.