Abstract: Using indole squaraine cyanine as the fluorescent scaffold, we successfully designed and synthesized a wash free fluorescent probe, designated as CSE, through the introduction of cationic salts. Performance characterization demonstrated that CSE not only exhibits excellent photostability but also possesses near-infrared fluorescent emission. Intracellular co-localization assays with commercial mitochondria-specific dyes further validated its ability to precisely target mitochondria. Under stimulated emission depletion (STED) super-resolution microscopy with depletion of 775nm, CSE clearly resolved the fine morphological details of mitochondrial cristae, achieving an impressive spatial resolution of up to 54 nm. During super-resolution dynamic tracking of mitochondria leveraging CSE, we successfully captured and delineated key dynamic events, including mitochondrial fusion and fission, cristae remodeling, and tubulation of mitochondria. The CSE probe developed herein enables super-resolution imaging and dynamic tracking of mitochondrial cristae, thereby holding great promise as a powerful tool for dissecting the dynamic mechanisms of mitochondrial cristae under physiological conditions. Keywords fluorescent probe; STED; mitochondrial cristae.

Key words: Fluorescent probe, Stimulated emission depletion(STED), Mitochondrial cristae