Pd 1 percp cy5

B‐cell subsets and cTfh were characterized using following surface markers: CD19‐FITC (Fluorescein isothiocyanate; Clone J4.119, Beckman Coulter, Indianapolis, IN, USA), CD38‐APC (Allophycocyanin; Clone HB‐7, BD Biosciences, San Jose, CA, USA), CD24‐PerCP‐Cy5.5 (Peridinin chlorophyll protein complex‐cyanine 5.5; Clone ML5, BD Biosciences), CD27‐PE (phycoerythrin; Clone L128, BD Biosciences), CD4‐PC7 (PE‐Cyanine7; Clone 13B8.2, Beckman Coulter), CXCR5‐ Alexa Fluor^®488 (Clone RF8B2, BD Biosciences), PD‐1‐PerCP‐Cy5.5 (Clone EH12.1, BD Biosciences), and ICOS‐APC (Clone ISA‐3, BD Biosciences). The gating strategy of B‐cell subsets is shown in Supplementary Figure 1, while that of cTfh is shown in Supplementary Figure 2.

Tumor xenografts were cut into pieces and digested with collagenase IV (Sigma, USA) and DNAse I (Sigma, USA) for 30 min at 37°C. Then, cell suspensions were passed through a 70-μm strainer to remove undigested tissues. Erythrocytes were removed by Red Blood Cell Lysis Solution (Qiagen, USA). For cell surface staining, 1 × 10⁶ cells were incubated with anti-Fc receptor blocking antibody at 4°C for 30 min. For intracellular staining, 1 × 10⁶ cells were fixed and permeabilized using the Fixation/Permeabilization Solution Kit (BD Bioscience, USA) according to the manufacturer’s instructions. Then, cells were stained with the indicated antibodies for 30 min at 4°C. Flow cytometry was performed using a BD Influx cell sorter (BD Bioscience). Flow cytometry data were analyzed by FlowJo version 10 (FlowJo, USA). The flow cytometry antibodies used in our study were CD45 APC, CD4 PE, CD8a V450, CD25 APC-CY7, FoxP3 PE-CY7, IFN-γ PE-CY7, IL-4 APC-CY7, Gr-1 V450, Ly6C FITC, CD11 b PE, F4/80 APC-CY7, Tim-3 PE-CY7, PD-1 PerCP CY5.5, T-bet FITC, GATA3 APC, TNF-α APC-CY7, anti-Ki67 FITC, and anti-Granzyme B PE, all from BD Bioscience.