alexa fluor 647 anti human cd39 antibody  (R&D Systems)

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: The roles of CD39⁺CD8⁺ TILs in the tumor immune microenvironment. The left panel depicts CD39⁺CD8⁺ T cells with features consistent with antitumor activity, including increased TCR clonal expansion and an effector-like feature. The right panel shows that, in bladder cancer patient cohorts, CD39⁺CD8⁺ TILs are associated with clinical outcomes. In parallel, adoptive transfer experiments in a murine bladder cancer model support a functional contribution of CD39⁺CD8⁺ T cells to tumor control. Overall, the scheme highlights CD39 as a practical biomarker to enrich tumor-reactive CD8⁺ TILs and to inform the development of cancer immunotherapy.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Activity Assay, Adoptive Transfer Assay, Functional Assay, Control, Biomarker Discovery

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: Clinical prognosis association and single-cell distribution of CD39 (ENTPD1). (A) Kaplan-Meier analysis of overall survival stratified by ENTPD1 expression across multiple solid tumor types in TCGA. (B) Kaplan-Meier analysis stratified by ENTPD1 expression in patients receiving immune checkpoint blockade. (C) t-SNE of integrated scRNA-seq data from 20 BLCA patients, annotated into 9 major cell types. (D) FeaturePlot of ENTPD1 expression across all cells and split by tissue origin (bladder normal vs tumor). (E) Average ENTPD1 expression across major cell types. (F) Comparison of average ENTPD1 expression between tumor and normal tissues within each major cell type. (G) t-SNE visualization of the CD8 + T cell populations from bladder tumor. (H) FeaturePlot of ENTPD1 expression across CD8⁺ T cell subsets in tumor tissue. (I) Average ENTPD1 expression across CD8⁺ T cell subsets.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Single Cell, Expressing, Comparison

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: Single-cell transcriptional profile and bulk TCR repertoire of human bladder cancer. (A) t-SNE visualization of integrated scRNA-seq data showing CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (B) Comparison of the cell ratio of major cell types in CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (C) Volcano plot shown the differentially expressed genes between CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (D) GO enrichment analysis of genes upregulated in CD39⁺CD8⁺ TILs. (E) GSEA enrichment plots of T cell function-related genes upregulated in CD39⁺CD8⁺ TILs. (F) Signature scores for tumor reactivity, tumor specificity, mutation-associated neoantigen (MANA) TIL, proliferation and virus-specific gene signatures. (G) Expression of CD39 + CD8 + TIL signature-related genes (G) Kaplan-Meier curves for overall survival stratified by CD39⁺CD8⁺ TIL signature enrichment in the TCGA-BLCA cohort (n = 424) and the IMvigor210 cohort (n = 195). (H) Tree maps of TCR clonotypes in CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (I) Relative frequencies of the top 10 most abundant TCR clonotypes. **** P < 0.0001.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Single Cell, Comparison, Cell Function Assay, Mutagenesis, Virus, Expressing

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: Tumor-infiltrating CD39⁺CD8⁺ T cells are enriched in BLCA, display an effector-like phenotype, and associate with tumor stage and patient survival. (A) Representative double-stained immunohistochemistry (IHC) image for CD39 (brown) and CD8 (pink) in tumor tissue slice. Black arrowheads indicate CD8 + T cell, red arrowheads indicate CD39 + CD8 + T cell. Scale bar, 20 μm. (B) Quantification of CD8⁺ T cell infiltration in tumor and normal bladder tissues. (C) Quantification of CD39⁺CD8⁺ T cell infiltration in tumor and normal bladder tissues. (D) Representative flow cytometry gating of CD8 expression of CD3 + T cells and quantification of CD8⁺ T cell frequencies across tumor stages. (E) Representative flow cytometry gating of CD39 expression of CD8 + T cells and quantification of CD39 + CD8⁺ T cell frequencies across tumor stages. (F) Correlation between CD39 expression and PD-1, CD103, CD134, and CD137 expression on tumor-infiltrating CD8⁺ T cell. (G) Flow cytometric comparison of CD103, CD134, and CD137 frequencies between CD8⁺CD39⁻ and CD8⁺CD39⁺ T cells. (H) Kaplan-Meier analysis of overall survival stratified by intratumoral CD8 + T cell and CD39⁺CD8⁺ T cell infiltration in the NJDT cohort (n = 82). ** P < 0.01, **** P < 0.0001.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Staining, Immunohistochemistry, Flow Cytometry, Expressing, Comparison

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: Single-cell transcriptomic profiling identifies an effector-like program in CD39⁺CD8⁺ TILs from the murine MB49 model. (A) Experimental workflow for FACS isolation of CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs from MB49 tumors followed by paired scRNA-seq and scTCR-seq. (B) t-SNE visualization of scRNA-seq data showing the distribution of CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (C) Comparison of the cell ratio of major cell types in CD39⁺CD8⁺ TILs and CD39⁻CD8⁺ TILs. (D) Volcano plot shows the differentially expressed genes between CD39 + CD8 + TILs and CD39 - CD8 + TILs. (E) GO enrichment analysis of genes upregulated in CD39⁺CD8⁺ TILs. (F) GSEA enrichment plots of T cell function-related genes upregulated in CD39⁺CD8⁺ TILs. (G) Signature scores for tumor reactivity, tumor specificity and virus-specific gene signatures. **** P < 0.0001.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Single Cell, Isolation, Comparison, Cell Function Assay, Virus

Journal: International Journal of Biological Sciences

Article Title: Clonotype-Resolved Single-Cell Multi-Omics Unlocks the Profile of Tumor-Infiltrating CD39⁺CD8⁺ T Cells and Enables Adoptive Cell Therapy for Solid Tumor

doi: 10.7150/ijbs.130389

Figure Lengend Snippet: Clonotype-resolved single-cell TCR analysis identifies tumor-reactive TCR sequence from CD39⁺CD8⁺ TILs. (A) Distribution of TCR clonotypes from MB49 tumor. (B) Stacked chart shows TCR clone-size distribution of CD39 +/- CD8 + T cells population. (C) Tree maps of TCR clonotypes in CD39 +/- CD8 + TIL population. (D) Co-expression of CD39 and N4 tetramer among CD8 + TILs in MB49-OVA tumor. (E) mRNA expression levels of Tnf , Ifng , Gzma , Gzmk , Prf1 and Nkg7 in the dominant TCR clonotype of CD39 + CD8 + TILs and the top 5 TCR clonotypes from CD39 - CD8 + TILs. (F) Scheme summarizes the activation and cytotoxicity assay of tumor reactive TCR. (G-H) Expression of CD69 on TCR-Jurkat cells after co-culture with MB49 tumor cells. (I) Viral infection efficiency of TCR-T cells. (J) Representative microscopy image after 24 hours co-culture of TCR-T cells with MB49 tumor cells. (K) Tumor cell killing by TCR-T cells in co-culture with MB49 cells. (L) TCR-Jurkat co-cultured with tumor cells of different MHC background, including CT26 (H-2K d ) or UMUC3 (HLA-A*02). (M) TCR-Jurkat co-cultured with normal renal cells. ns, P > 0.05, *** P < 0.001.

Article Snippet: For double IHC staining, anti-human CD8 monoclonal antibody (Proteintech, cat#66868-1-Ig) and anti-human CD39 polyclonal antibody (Proteintech, cat#14211-1-AP) were used for IHC staining.

Techniques: Single Cell, Sequencing, Expressing, Activation Assay, Cytotoxicity Assay, Co-Culture Assay, Infection, Microscopy, Cell Culture