Fig. 4: In vitro cellular investigation.

Representative CLSM images of (a) 4T1 and (b) RAW cells upon incubation with BC@Z or BC@Z-M for 4 h. The cell nuclei were stained with 4’,6-diamidino-2-phenylindole (DAPI; blue fluorescence). Scale bars: 50 μm. c Quantitative data showing the mean fluorescence intensity (MFI) based on (a, b) (n = 4 independent samples). d NIR-I and NIR-II microscopic fluorescence images of 4T1 cells after incubation with BC@Z-M for 0 or 3 h under hypoxic condition. Scale bars: 50 μm. e MFI of 4T1 cells treated with BC@Z-M under normoxic or hypoxic conditions for 0 or 3 h (n = 3 independent samples). f NIR-I and NIR-II IVIS images of 4T1 cells cultured with BC@Z-M in 12-well plates under normoxic or hypoxic conditions for 3 h based on (d). g Normalized ratiometric FL of 4T1 cells treated with BC@Z-M under hypoxic or normoxic conditions based on (f) (n = 3 independent samples). h Representative CLSM images showing the ROS generation in 4T1 cancer cells after different treatments as indicated. Scale bar: 50 µm. i Cell viability of 4T1 tumor cells was assessed using MTT assay following treatment with varying concentrations of BC@Z-M under hypoxic conditions with or without 730 nm light irradiation (n = 4 independent samples). j Representative FL microscopic images showing 4T1 tumor cells co-stained with calcein-AM and propidium iodide (PI) following different indicated treatments. Scale bar: 100 μm. All data are presented as mean ± SD. For (a, b, d, h, j), experiment was repeated three times independently with similar results. For (c, e, g, i), statistical significance was determined using two-tailed Student’s t-test. Source data are provided as a Source Data file.