Mitochondria-Immobilized Fluorescent Probe for the Detection of Hypochlorite in Living Cells, Tissues, and Zebrafishes.

A mitochondria targeting and immobilized fluorescent probe (Rd1) using triphenylphosphonium as the targeting group and methoxymaleimide as the fixed site is designed for the detection of ClO-. The methoxymaleimide fixed group can react with nucleophiles, such as the reactive thiol groups present in mitochondrial polypeptides and proteins, and form covalent bonds to immobilize the probe within mitochondria. The immobilization of Rd1 enhances its ability to withstand the risk of leakage from mitochondria. Methoxymaleimide shows better reactivity toward Cys than glutathione (GSH), which decreases the ineffective labeling of GSH when it covalently bonds with the reactive thiol residues of mitochondrial proteins; furthermore, it can resist hydrolysis during a long-term storage in water, compared with the classic benzyl chloride fixed unit. The imaging results indicate that Rd1 displays enhanced retention within the mitochondria of cells and tissues upon the decrease of mitochondrial membrane potential (MMP) caused by different stimulations. Furthermore, it possesses the ability to visualize exogenous and endogenous ClO- in living cells, tissues, and zebrafishes.