Anti mouse cd45 brilliant violet 421

The harvested tumors of the mice were cut into pieces in DMEM medium containing 2% FBS on ice and then treated with 5 volume equivalent trypsin at 37℃ for 60 min. The samples were filtered and centrifuged at 600 g for 5 min. The cells were washed with PBS containing 2% FBS and co-incubated with sterile ZombieNIR dye and Fc receptor blocker at 4℃ for 15 min. Next, the cells were stained with the primary antibodies in dark for 30 min, including anti-mouse CD86-PE antibody (B334834, Biolegend), anti-mouse CD206-APC (B354282, Biolegend), anti-mouse F4/80-PE/Cyanine-7 (B342137, Biolegend), anti-mouse CD45-Brilliant Violet 421 (B343559, Biolegend), anti-mouse CD3-PE (B341466, Biolegend), anti-mouse CD4-FITC (B269033, Biolegend), anti-mouse CD8a-APC (B348048, Biolegend), anti-mouse CD11c-APC (B339313, Biolegend), anti-mouse I-A/I-E (MHC II)-PE/Cyanine-7 (B337001, Biolegend) and anti-mouse CD11b-FITC (B349919, Biolegend). After rinsing excess antibodies, the cells were immediately assessed by a flow cytometer (Beckman Coulter, USA).

The following antibodies were used: anti-mouse CD3e-FITC, -PerCP-Cy5.5, or -APC-Cy7 (clone 145-2C11; BioLegend), CD4-APC or -PE-Cy7 (clone RM4-5; BioLegend), anti-mouse CD8α-Alexa Fluor 488 (clone 53-6.7; BioLegend), anti-mouse CD8β-PE (clone YTS156.7.7; BioLegend), anti-mouse CD27-Brilliant Violet 510 (clone LG.3A10; BioLegend), anti-mouse CD28-Brilliant Violet 421 (clone 37.51; BioLegend), anti-mouse CD45- Brilliant Violet 421 (clone 30-F11; BioLegend), anti-mouse CD274 (PD-L1)-PE (clone 10F.9G2; BioLegend), anti-mouse epithelial cell adhesion molecule (EpCAM)-APC (clone G8.8; BioLegend), anti-mouse MHC-II-FITC (clone M5/114.15.2; eBioscience), anti-mouse TCRβ- APC-Cy7 (clone H57-597; BioLegend) or anti-mouse γδTCR -PE-Cy7 (clone GL3; BioLegend). For scRNAseq analysis, anti-mouse Hashtags 1 to 4 (M1/42; 39-F11, Biolegend, 155861, 155863, 155865, 155867) were used.

The following flow antibodies were purchase from Biolegend: Antimouse CD45-Brilliant Violet 421; Antimouse Ly-6G-Alexa Fluor 488; Antimouse Ly-6G-Alexa Fluor 647; Antimouse CD31-Alexa Fluor 647; Antimouse CD31-PE/Cyanine7; Antimouse CD45 FITC; Anti-mouse/human CD11b-Brilliant Violet 421; Antimouse CD184 (CXCR4)-PerCP/Cyanine5.5; Antimouse CD182 (CXCR2)-APC/Cyanine7.
The following flow antibodies were purchase from Invitrogen: Anti-CD144 (VE-cadherin)-eFluor 660; Anti-CD54 (ICAM-1)-PE; Anti-CD11b-APC; Antimouse F4/80-PE.
The following flow antibodies were purchase from BD Biosciences: Antimouse Ly6G-PE; Anti-Mouse CD16/CD32. Anti-Fibrin (Cat#MABS2155) was purchased from Sigma-Aldrich; Anti-SOD2/MnSOD (Cat#ab68155) and Anti-CD15 (Cat#ab135377) were was purchased from Abcam; Antirabbit β-Actin-HRP (Cat#AF5006) was purchased from Beyotime; Antirabbit IgG-HRP (Cat#7074 S), anti-Ly6G (Cat#87048 S) and anti-COX IV (Cat#4844 S) were purchased from Cell Signaling Technology; Anti-TOMM22 (Cat#66562-1-Ig) was purchased from Proteintech; InVivoMAb antimouse Ly6G (Cat#BE0075-1) and InVivoMAb rat IgG2a isotype control (Cat#BE0089) were purchase from Bio X Cell; Donkey antirabbit IgG (H + L)-Cy3 (Cat#711-165-152) and donkey anti-mouse IgG (H + L)-Cy3 (Cat#715-165-150) were purchased from Jackson ImmunoResearch.