Optimizing SiQDs-(K/L-P)ox Incubation Time for ONOO- Detection and Co-localization in HepG2 Cells

Additionally, the phagocytic capacity of HepG2 cells was investigated through the formative process of SiQDs-(K/L-P)ox colocation and detecting ONOO- in HepG2 cells, as depicted in Figure 5a. The (K/L-P)ox, a biological macromolecule synthesized by peptides, normally cannot cross the cell membrane. The synthesized SiQDs-(K/L-P)ox has an ultra-small particle size of 4 nm and combines hydrophilic groups (-COOH, -OH and -NH3 etc.) and hydrophobic groups (benzene ring) on its surface. This allows it to freely enter the phospholipid bilayer through diffusion, enabling ONOO- detection and co-localization.

To investigate the phagocytic capacity of HepG2 cells, SiQDs-(K/L-P)ox (50 μg/mL) and cells were cultured at 37 °C under 5% CO2 for 5, 10, 15, 30, 60 and 120 min. Confocal microscopy showed that SiQDs-(K/L-P)ox emitted green and yellow fluorescence under the excitation of a 488 and 514 nm laser. The 514 nm channel was chosen for the next study as the yellow fluorescence was stronger. The fluorescence intensity increased with the incubation time, reaching the maximum fluorescence intensity at 120 min (75.7) as shown in Figure S11 and Table S1. However, excessive uptake of QDs by cells resulted in high fluorescence intensity and blurred cell contour. After co-incubation for 30 min, the fluorescence intensity of HepG2 cells was 62.6, with clear cell morphology. Thus, the incubation time of the probes with the cells in subsequent biological experiments was set to 30 min.