The increasing industrial use of palladium has led to its environmental accumulation, raising concerns about its toxicity to aquatic life and human health. Therefore, fluorescent probes capable of detecting Pd2+ are highly beneficial. With this objective, a new near-infrared (NIR) fluorescent probe based on a cyanine dye, 2 ((E)-2-((E)-2-((dimethylcarbamothioyl)oxy)-3-(2-((Z)-1,3,3-trimethylindolin-2ylidene)ethylidene)cyclohex −1-en-1-yl)vinyl)-1,3,3-trimethyl-3H-indol-1-ium iodide (CNS), was synthesized for selective and rapid detection of Pd2+. The detection reaction followed the elimination of thiocarbamate moiety, leading to the highly fluorescent product. CNS demonstrated remarkable sensitivity (detection limit: 0.105 μM), high selectivity, short response time (1.0 min), long lifetime (0.88 ns), and easily detectable color changes upon Pd2+ exposure. A CNS-loaded TLC strip integrated with a smartphone detection system was able to detect Pd2+ in solutions, soil, and drug samples. In addition, CNS enabled concentration-dependent detection of Pd2+ in onion roots and epidermis. Because of low cytotoxicity, good membrane permeability, NIR fluorescence, and high contrast, CNS has been successfully applied to Pd2+ bioimaging in living cells, targeting mitochondria. Compared to existing probes, CNS offers superior sensitivity, selectivity, and adaptability for sensing applications.