Integrated fluorescence microscopic imaging system

The TEM images were recorded on a JEOL JEM-1200EX (Japanese Electronics and America GATAN). The DLS and Zeta potentials were obtained by NanoBrook Omni particle sizer (Brookhaven Instruments, USA). The XRD pattern was recorded on a D8 ADVANCE diffractometer (Brucker, Germany). The UV–Vis absorption curve was obtained using a UV-2550 Spectrophotometer (Shimadzu, Kyoto, Japan). Fourier transform infrared spectroscopy (FTIR) spectra were collected on a Nicolet IS10 (American Thermoelectric). The XPS were measured using a Thermo Escalab 250Xi (Thermo Scientific Escalab, USA). Specific surface area and corresponding pore-size distribution were determined on a Micromeritics Tristar 3000 system (ASAP, 2460, Micromeritics, USA) and tested by the nitrogen (N₂) adsorption–desorption isothermal method. CCK-8 content were recorded using a microplate reader. The fluorescence images of Live and Dead, intracellular reactive oxygen species (ROS) and were obtained by an integrated fluorescence microscopic imaging system (Keyence, China). Cell uptake image and EdU-488 cell proliferation assay were observed by confocal laser scanning microscopy (CLSM).

Caco-2 cells were cultured in 6-well plates until reaching about 80% confluence. The cells were treated with different experimental groups including HPB, CaO₂, DOX, PCDP, and PCDP + NIR, while samples treated with PBS were utilized as controls. The cells of PCDP + NIR group was additionally irradiated with an 808 nm laser for 5 min after incubating PCDP and cells together for 12 h. After incubating for 24 h, the cells were stained with acridine orange/ethidium bromide (AO/EB) for 5 min and observed under an integrated fluorescence microscopic imaging system (Keyence, China). Green and red fluorescence indicated live and dead cells, respectively.