Percp cyanine5.5 ly6g

The lipids 18: 1TAP (1, 2-dioleoyl-3-trimethylammonium-propane), DOPE (1, 2-dioleoyl-sn-glycero-3-phosphoethanolamine), EPC, 16:0 NBD PE, and 16:0 Liss Rhod PE were purchased from Avanti Polar Lipids, Inc. (Alabaster, AL, USA). DSPE-PEG2000-Folic acid was purchased from Nanocs (Boston, MA, USA). Chol (Cholesterol), Collagenase, and DNase I were purchased from Sigma-Aldrich (St. Louis, MO, USA).
The dye DiI was purchased from Beyotime Biotechnology. PTX was purchased from Meilunbio^® (Dalian, China). Paclitaxel injection was purchased from HAPHARM Group Co., Ltd. (Harbin, China). The CD9, CD63, and TSG101 antibodies were purchased from Abcam (Cambridge, MA, USA).
The antibodies of flow cytometry (anti-mouse): PE-CD45, FITC-CD4, APC-CD8a, PerCP/Cyanine5.5-CD69, FITC-CD3ε, APC-CD49b (pan-NK cells), APC/Cyanine7-CD335 (NKp46), PE-CD107a (LAMP-1), APC-CD45R/B220, PE/Cyanine7-CD69, FITC-CD45, APC/Cyanine7-F4/80, PE-CD11c, PE/Cyanine7-CD86, APC anti-mouse/human CD11b, PerCP/Cyanine5.5-Ly-6G, FITC-Ly-6C, PE/Cyanine7-CD45, APC-CD25, PE-FOXP3, PE-CD86, and PerCP/Cyanine5.5-CD206 (MMR) were purchased from Biolegend (San Diego, CA, USA).

BALF was centrifuged at 300g for 5 min and 4 ℃. The supernatant was stored at -80 °C. Neutrophils were identified as CD45⁺Ly6G⁺CD11b⁺ when the cell sediments were resuspended in PBS and stained with FITC-anti-CD45, PerCP/Cyanine5.5-Ly6G, and APC/Cyanine7-CD11b (BioLegend, San Diego, CA, USA) 27 (link). Total cells in the BALF were counted using a hemocytometer.

Lymphocytes isolated from livers by percoll (Sigma, St. Louis, MO, USA) were washed three times in phosphate-buffered saline (PBS) containing 1% fetal bovine serum (FBS; FACS Buffer). For detecting dead cells, harvested cells were stained via a LIVE/DEAD Fixable Far-Red-Dead Cell Stain Kit (1:1000) using either 633 or 635 nm excitation (Cat#L34973, Thermo Fisher Scientific). For Brilliant Violet 510-CD45 (Cat#103137, BioLegend) (1:400), Alexa Fluor 700-CD11b (Cat#101222, BioLegend) (1:400), PerCP/Cyanine5.5-Ly6G (Cat#127616, BioLegend) (1:600), and PE/Cyanine7-F4/80 (Cat#123114, BioLegend) (1:400) staining, harvested cells were washed and incubated in PBS containing 1% FBS containing fluorochrome-conjugated antibodies in a U-bottom 96-well plate. The frequency of the different cell population was measured via flow cytometry (BD LSRFortessa, BD Biosciences) and analyzed by FlowJo_v10 (BD, Ashland, OR, USA). Antibodies used in this study for FC were listed in Supplementary Materials.