Avanti mini extruder

The different cancer cell-derived ApoBds were produced by serum starvation, as described previously.^{43 (link)} Briefly, the different cancer cells (human breast cancer cells (SKBR3, MCF-7, and MDA-MB-231), human glioblastoma cells (U87-MG and LN229), human hepatocellular carcinoma cells (HepG2), and mouse 4T1 breast cancer cells were plated to 80% confluence (4 × 10⁶ cells/10 cm plate) for 24 h. They were then washed three times with PBS. We added 8 mL of serum-free DMEM with Pen/Strep for 72 h, and the cell apoptosis process was assessed by fluorescence microscopy and flow cytometry using Hoechst 33342 and FITC-annexin V staining. The cell free culture medium was collected for the isolation of tumor cell-derived ApoBds (ApoBds), as shown in Figure S1 and as mentioned earlier.^{19 (link)} Briefly, the cell-free medium was collected and centrifuged at 300g for 5 min to remove cells and large debris. The supernatant was further centrifuged again at 2000g for 30 min to collect the apoptotic bodies, and the pellet was carefully resuspended in 300 μL of PBS. The isolated ApoBds from different cancer cells were mildly sonicated to break them into small proteolipid vesicles and then reconstructed with or without vancomycin (freeze and thaw) by a physical extrusion process using an Avanti mini extruder (Avanti Polar Lipids Inc.), as mentioned earlier.^{19 (link)}