1：Experimental Design and Method Selection:

The study involved designing and synthesizing a naphthalene-indoline-based fluorescent probe (1-Red) that targets lysosomes and becomes fluorescent upon oxidation by ?OH. Two-photon microscopy was used for imaging due to its advantages in biological applications.

2：Sample Selection and Data Sources:

Mouse embryonic fibroblast (MEF) cells were used as the experimental model, cultured under standard conditions.

3：List of Experimental Equipment and Materials:

Equipment included a Bruker Advance III Ascend 400 MHz NMR spectrometer, Thermo Fisher Orbitrap mass spectrometer, JASCO FT/IR-4100, Cary 100 Bio UV-Vis Spectrophotometer, JASCO FP-750 Spectrofluorometer, Leica SP5 MP confocal microscope with MaiTai Ti:Sapphire HP laser, and various chemicals and reagents (e.g., isobutyryl chloride, phenylhydrazine hydrochloride, sodium borohydride).

4：Experimental Procedures and Operational Workflow:

Synthesis of compounds 2, 3, 1, and 1-Red through Friedel-Crafts, Fisher indole reactions, methylation, and reduction. Cell culture and incubation with probes, followed by two-photon microscopy imaging. Selectivity tests with various ROS and other species.

5：Data Analysis Methods:

Fluorescence intensity measurements, NMR spectroscopy, mass spectrometry, and statistical analysis including Pearson correlation coefficient for colocalization studies.