Lysotracker green dnd 26

The cells on the coverslips after clinorotation were incubated in media containing 250 nmol/L MitoTracker Deep Red FM (40743ES50, Yeasen, Shanghai, China) for 30 min at 37°C and washed with pre-warmed PBS. The cells were then incubated in media containing 50 nmol/L LysoTracker Green DND-26(40738ES50, Yeasen) for 1 h in the cell incubator and observed under a confocal microscope (LSM 800, Zeiss, Oberkochen, Germany) to determine the red and green fluorescence.

Mesoporous silica was purchased from nanoComposix (USA). Shanzhiside methylester was purchased from DESITE (China). DOX and Dialysis Membrane (2 kD) were purchased from Solarbio (China). Cy5, Hoechst 33342, and LysoTracker Green DND-26 were purchased from Yeasen Biotechnology (China). DSPE-PEG-FITC, MW:2000 purchased from Ruixi Biology Co., Ltd (China). Annexin V-FITC apoptosis detection kit was purchased from Beyotime Biotechnology (China). ROS assay kit was purchased from Beyotime Biotechnology (China). JC-1 MitoMP Detection Kit and cell counting kit-8 (CCK-8) were purchased from Dojindo Laboratories (Japan). Penicillin and streptomycin cocktail, fetal bovine serum (FBS), and RPMI-1640 were purchased from Life Technologies (USA). Anti-Bcl-2, anti-Bax, anti-TNF-α and anti-IL-1β, anti-CD47, and anti-SIRPα were manufactured by Cell Signaling Technology (USA). Anti-Ly6c was purchased from Abcam (UK). TNF-α ELISA. IL-1β ELISA was purchased from R&D Systems (China). HRP conjugated goat anti-rabbit IgG and HRP conjugated goat anti-mouse IgG were manufactured by Auragene Biotech (China). TdT in situ apoptotic kit was purchased from R&D Systems (China). Hematoxylin eosin (H&E) and DAPI were purchased from Servicebio Technology (China). Polycarbonate porous membrane syringe filters (200 nm) were provided by Whatman (USA).

During the in vitro transcription of sPD-L1 mRNA, 25% Cy3-UTP (B8330, APE×BIO) was incorporated. Subsequently, the fluorescent mRNA was encapsulated within DOTAP-LNPs. After 18 hours of LNPs transfection in cells, LysoTracker Green DND-26 (40738ES50, Yeasen) was added and incubated for 30 minutes. Live cell imaging was then performed using a Leica DMi8 S Live cell microscope.