Abstract: Lysosomes are intracellular organelles with lysosomal or digestive functions which contain various acid-dependent enzymes that are responsible for the recycling and degradation of intracellular substances. pH changes in lysosomes can reduce the activity of various enzymes, impairing the cell′s regulatory capacity and ultimately leading to apoptosis or pyroptosis. Therefore, monitoring lysosomal pH values is valuable for investigating alterations in intracellular physiological and pathological processes. This study developed a novel pH-responsive near-infrared (NIR) fluorescent probe targeting lysosomes, named piperazine-croconic acid-piperazine (PCP). This probe was formed by connecting the electron-donating group piperazine to both ends of croconic acid. The π-conjugated system, featuring a five-membered ring structure, endowed the probe with excellent stability and resistance to photobleaching. The tertiary amine moiety of piperazine, serving as a lysosome-targeting group, produced reversible protonation/deprotonation under varying pH conditions as result of exhibiting pH-responsive characteristics. The synthesized PCP demonstrated excellent NIR fluorescence performance, making it suitable for fluorescence imaging of lysosomal pH values in living cells. Both solution and intracellular experiments confirmed that PCP exhibited good biocompatibility and pH-responsive characteristics. Additionally, its symmetrical structure and molecular rigidity contributed to its excellent resistance to photobleaching. This study employed PCP probes to detect the pH values of the lysosomes, offering potential avenues for the treatment and prevention of diseases associated with lysosomal dysfunction.