Understanding the pathological process of biological systems can greatly improve the prevention and treatment of diseases. The study of pathological processes has now reached the molecular level, and molecular fluorescent probes have become a powerful tool. Chromene, also known as benzo-pyran molecule, is a structural element of natural products with good biological compatibility and was developed as a fluorescent probe. The thiol-chromene "click" nucleophilic pyran ring-opening reaction allows the quick detection of thiol. In this work, the chromene alcohol can function as an efficient self-immolative spacer, which covalently links NIR fluorophore via a carbonyl ester. Due to its favorable characteristics and superior applicability, the self-immolative amplifier NIR-HMPC achieves the specific, rapid, sensitive, NIR fluorescent detection of thiols. Furthermore, the indoles iodized salt in the system can specifically target thiols in mitochondria. Thus, this probe was used to visualize the fluctuations of thiols during oxidative stress and cell apoptosis, cerebral ischemia reperfusion, demonstrating that it is valuable for elucidating pathophysiology process in living organism. This discovery provides an effective means for studying the pathological process of thiol related diseases.