anti mouse cd8 mabs  (Bio X Cell)

Journal: Nature Communications

Article Title: Histone lactylation increases CXCL1 expression for neutrophil infiltration and immune escape in pancreatic cancer

doi: 10.1038/s41467-026-69311-5

Figure Lengend Snippet: A The expression of neutrophils and CD8 + T cells in low- and high-glycolysis clusters, divided by ssGSEA score of glycolysis-related genes, in TCGA and CPTAC PAAD database. Data are presented as box plots showing the median (center line), the first and third quartiles (box bounds), and the whiskers extend to 1.5 times the interquartile range from the box. B The correlation of glycolysis level and neutrophils or CD8 + T cells expression in TCGA ( n = 179 patients) and CPTAC ( n = 140 patients) PAAD database. C Kaplan–Meier plots representing survival probabilities in TCGA-PAAD patients according to the relative level of glycolysis-related or neutrophil-related gene expression. D The diagram illustrating the targets of 2DG and shRNA in the glycolysis process. E CyTOF analysis of tumor-infiltrating immunocytes in subcutaneous tumor models with or without 2DG treatment: tSNE plots showing 12 meta-clusters based on the expression of 41 markers for the immunocytes. F Bar chart of the frequencies of the immune cell subsets in two experimental groups. G A schematic diagram showing the orthotopic PDAC model ( n = 6 mice per group in one experiment) with or without 2DG treatment. H , I Image and weights of the orthotopic tumors in the experimental groups from ( G ) at the end of the experiments. ( n = 6 mice per group). ( J – M ) Tumor-infiltrating neutrophils, CD8 + T cells, GZMB + CD8 + T cells, and PD-1 + CD8 + T cells isolated from the orthotopic tumors were analyzed using flow cytometry: representative graphs of flow cytometry (left panel) and statistical analysis of the cell ratio (right panel) ( n = 6 mice per group). N A schematic diagram showing the orthotopic tumor model in C57BL/6 J mice ( n = 6 mice per group in one experiment) treated with or without anti-Ly6G/anti-CD8 antibody. O , P Image and weights of the orthotopic tumors in the experimental groups from ( N ) at the end of the experiments ( n = 6 mice per group). Q – S Tumor-infiltrating neutrophils, CD8 + T cells, and GZMB + CD8 + T cells isolated from the orthotopic tumors were analyzed using flow cytometry: Representative graphs of flow cytometry (left panel) and statistical analysis of the cell ratio (right panel) ( n = 6 mice per group). T , U Representative luminescence images of the mouse model in ( N ) and statistical analysis of the total flux ( n = 6 mice per group). Data represent mean ± SEM. Statistical analysis was conducted using the two-tailed unpaired Student’s t test ( A, I–M, P–S, and U ), two-tailed Pearson’s correlation analysis ( B ), and log-rank test ( C ). Source data are provided as a Source Data file.

Article Snippet: The antibodies used here included anti-L-Lactyl-Histone H3K18 antibody (1:500, PTMBIO, #PTM-1406RM), anti-GRO antibody (1:200, AFFINITY, #AF5403), Ki-67 (1:400, Servicebio, # GB111499 ), anti-Ly6G antibody (1:500, Servicebio, #GB11229), and anti-CD8 antibody (1:200, Cell Signaling Technology, #70306), and anti-PCAF antibody(1:200, Cell Signaling Technology, #3378).

Techniques: Expressing, Gene Expression, shRNA, Isolation, Flow Cytometry, Two Tailed Test

Journal: Nature Communications

Article Title: Histone lactylation increases CXCL1 expression for neutrophil infiltration and immune escape in pancreatic cancer

doi: 10.1038/s41467-026-69311-5

Figure Lengend Snippet: A The flowchart illustrates that RNA sequencing was conducted using two PDAC cell lines with or without 2DG treatment. Subsequently, the intersection of detected genes within the chemokine family in both cell lines is identified, and a heatmap is generated to display the log 2 fold change (2DG versus vehicle) of these genes. B , C Relative mRNA levels of Cxcl1 in PDAC cell lines following glycolysis inhibition, either by using 2DG or silencing LDH, were analyzed by qRT-PCR ( n = 3 independent experiments). D , E Relative protein levels of CXCL1 in PDAC cell lines following glycolysis inhibition, either by using 2DG or silencing LDH, were determined using ELISA assay ( n = 3 independent experiments). F Representative IHC images of subcutaneous tumors ( n = 6 mice for 2DG treatment, n = 5 mice for shLDH/shNTC groups) stained by CXCL1 antibody (scale bars = 100 μm). G Relative serum CXCL1 levels in mice treated with or without 2DG were measured by ELISA ( n = 6 mice per group). H Relative serum CXCL1 levels in healthy donors and PDAC patients were measured by ELISA (22 healthy samples and 27 PDAC samples). I Schematic diagram showing the in vitro migration assay: human/mouse neutrophils were co-incubated with the culture medium supernatant from PDAC cells treated with 2DG or LDH-knockdown. J Neutrophils were co-cultured with CD8 + T cells in different proportions, and the proliferation of CD8 + T cells was detected with the CFSE assay ( n = 6 biologically independent samples). K The migratory activity of neutrophils co-incubated with the culture medium supernatant from PDAC cells treated with shLDH and recombinant CXCL1 was analyzed by counting the penetrated cell numbers ( n = 3 biologically independent samples). L The neutrophils were treated with SX-682 or Navarixin for 1.5 h in advance, and the migratory activity of neutrophils co-incubated with the culture medium supernatant from PDAC cell lines treated with shLDH was analyzed by counting the penetrated cell numbers ( n = 3 biologically independent samples). M Representative luminescence images of the orthotopic tumor in mice and statistical analysis of MFI ( n = 5 mice per group). N Tumor-infiltrating neutrophils isolated from the orthotopic tumors were analyzed using flow cytometry ( n = 5 mice per group). Data represent mean ± SEM. Statistical analysis was conducted using the two-tailed unpaired Student’s t test. Source data are provided as a Source Data file.

Article Snippet: The antibodies used here included anti-L-Lactyl-Histone H3K18 antibody (1:500, PTMBIO, #PTM-1406RM), anti-GRO antibody (1:200, AFFINITY, #AF5403), Ki-67 (1:400, Servicebio, # GB111499 ), anti-Ly6G antibody (1:500, Servicebio, #GB11229), and anti-CD8 antibody (1:200, Cell Signaling Technology, #70306), and anti-PCAF antibody(1:200, Cell Signaling Technology, #3378).

Techniques: RNA Sequencing, Generated, Inhibition, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Staining, In Vitro, Migration, Incubation, Knockdown, Cell Culture, CFSE Assay, Activity Assay, Recombinant, Isolation, Flow Cytometry, Two Tailed Test

Journal: Nature Communications

Article Title: Histone lactylation increases CXCL1 expression for neutrophil infiltration and immune escape in pancreatic cancer

doi: 10.1038/s41467-026-69311-5

Figure Lengend Snippet: A Molecular docking simulated the binding affinities of PCAF for acetyl-CoA and lactyl-CoA. B Co-IP was performed to confirm the interaction between H3K18la and PCAF in PANC1 and KPC cells. PCAF and H3K18la were detected on the same gel. C, D Western blot analysis demonstrates the level of H3K18la in PDAC cells with or without the treatment of PCAF inhibitors (bromosporine or Embelin). Histone H3 and H3K18la blots are from parallel-processed separate gels (size conflict) with samples from the same experiment. E Western blot analysis demonstrates the level of H3K18la in PDAC cells following the knockdown of Pcaf . Histone H3, PCAF, β-actin and H3K18la blots are from parallel-processed separate gels with samples from the same experiment. F Western blots of in vitro histone acetylation or lactylation assay, the samples as indicated. Blots are from parallel-processed separate gels with samples from the same experiment. G , H Relative Cxcl1 RNA and protein levels with the vehicle or bromosporine treatment were analyzed by qRT-PCR and ELISA ( n = 3 independent experiments), respectively. I Relative Cxcl1 RNA levels after Pcaf knockdown were analyzed by qRT-PCR ( n = 3 independent experiments). J A schematic diagram showing the orthotopic tumor construction in C57BL/6 J mice ( n = 5 mice per group in one experiment, 2 × 10 6 KPC-luc cells per mouse) with or without bromosporine treatment. K , L Image and weights of the orthotopic tumors in the experimental groups from ( J ) at the end of the experiments ( n = 5 mice per group). M Lactylation levels of H3K18 and PanK in orthotopic tumors with or without bromosporine treatment were detected by western blot ( n = 4 biologically independent samples per group). Histone H3, H3K18la, and PanKla blots are from parallel-processed separate gels (size conflict) with samples from the same experiment. N Serum CXCL1 levels in mice with or without bromosporine treatment were measured by ELISA ( n = 5 mice per group). O – R Tumor-infiltrating neutrophils, CD8 + T cells, GZMB + CD8 + T cells, and PD-1 + CD8 + T cells isolated from the orthotopic tumors were analyzed using flow cytometry. Representative graphs of flow cytometry (left panel) and statistical analysis of the cell ratio (right panel) ( n = 5 mice per group). S The migratory abilities of neutrophils co-incubated with the culture medium supernatant from shNTC/ Pcaf PDAC cells treated with recombinant CXCL1 were analyzed by counting the penetrated cell numbers ( n = 3 biologically independent samples). T Representative IHC images stained by CXCL1 or H3K18la antibody (scale bar = 100 μm, left panel), correlation analysis of H-scores of CXCL1 and H3K18la in PDAC samples ( n = 21 patients, right panel). Data represent mean ± SEM. Statistical analysis was conducted using the two-tailed unpaired Student’s t test ( G – I , L , and N – S ) and two-tailed Pearson’s correlation analysis ( T ). Unless otherwise indicated, all western blots had three independent experimental repetitions with consistent results. Source data are provided as a Source Data file.

Article Snippet: The antibodies used here included anti-L-Lactyl-Histone H3K18 antibody (1:500, PTMBIO, #PTM-1406RM), anti-GRO antibody (1:200, AFFINITY, #AF5403), Ki-67 (1:400, Servicebio, # GB111499 ), anti-Ly6G antibody (1:500, Servicebio, #GB11229), and anti-CD8 antibody (1:200, Cell Signaling Technology, #70306), and anti-PCAF antibody(1:200, Cell Signaling Technology, #3378).

Techniques: Binding Assay, Co-Immunoprecipitation Assay, Western Blot, Knockdown, In Vitro, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Isolation, Flow Cytometry, Incubation, Recombinant, Staining, Two Tailed Test

Journal: Nature Communications

Article Title: Histone lactylation increases CXCL1 expression for neutrophil infiltration and immune escape in pancreatic cancer

doi: 10.1038/s41467-026-69311-5

Figure Lengend Snippet: A A schematic diagram showing the subcutaneous tumor model ( n = 6 mice per group in one experiment, 6 × 10 6 PANC02 cells per mouse) treated with bromosporine and anti-PD-1 antibody. B. The tumor volume growth curves of subcutaneous tumors from ( A ). C , D Image and weights of the subcutaneous tumors at the end point of experiments ( n = 6 mice per group). E The statistical analysis of the cell ratio of tumor-infiltrating neutrophils and CD8 + T cells isolated from subcutaneous tumors ( n = 3 mice per group). F Western blot analysis demonstrates H3K18la levels in the subcutaneous tumors ( n = 3 biologically independent samples per group) from ( A ). Histone H3 and H3K18la blots are from parallel-processed separate gels (size conflict) with samples from the same experiment. G Relative serum CXCL1 levels in mice treated with or without bromosporine/anti-PD-1 antibody were measured by ELISA ( n = 6 mice per group). H A schematic diagram showing the combinational treatment schedule for the orthotopic KPC-luc tumor model ( n = 21 mice per group in one experiment, 2 × 10 6 KPC-luc cells per mouse). I , J Image and weights of the orthotopic tumors at the end point of experiments ( n = 6 mice per group). K – N Tumor-infiltrating neutrophils, CD8 + T cells, GZMB + CD8 + T cells, and PD-1 + CD8 + T cells were analyzed using flow cytometry. Representative graphs of flow cytometry (left panel) and statistical analysis of the cell ratio (right panel) ( n = 6 mice per group). O Western blot analysis showing the H3K18la levels in the orthotopic tumors ( n = 3 biologically independent samples per group) from ( H ). Histone H3 and H3K18la blots are from parallel-processed separate gels (size conflict) with samples from the same experiment. P Representative luminescence images and Quantification of radiance intensity of the mouse model in ( H ). Q Survival probability of mice with orthotopically transplanted PDAC ( n = 15 mice per group). R A working model displaying the signaling pathway through which the aerobic glycolysis-mediated Lactate-PCAF-H3K18la-CXCL1 axis modulates the tumor microenvironment in pancreatic cancer, and the scientific basis for the development of a novel therapeutic strategy for PDAC. Data represent mean ± SEM. Statistical analysis was conducted using the two-tailed unpaired Student’s t test ( B , D , E , G , J – N , P ) and the log-rank test ( Q ). Source data are provided as a Source Data file.

Article Snippet: The antibodies used here included anti-L-Lactyl-Histone H3K18 antibody (1:500, PTMBIO, #PTM-1406RM), anti-GRO antibody (1:200, AFFINITY, #AF5403), Ki-67 (1:400, Servicebio, # GB111499 ), anti-Ly6G antibody (1:500, Servicebio, #GB11229), and anti-CD8 antibody (1:200, Cell Signaling Technology, #70306), and anti-PCAF antibody(1:200, Cell Signaling Technology, #3378).

Techniques: Isolation, Western Blot, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Two Tailed Test

Journal: Journal of Virology

Article Title: Recombinant duck enteritis virus harboring the hemagglutinin genes of influenza virus rapidly induces specific cellular immunity in ducks

doi: 10.1128/jvi.02014-25

Figure Lengend Snippet: Expansion of T cells induced by rDEV-dH5/H7 and vDEV. Four-week-old ducks were inoculated i.m. with two doses of 10 5 TCID 50 of rDEV-dH5/H7 or vDEV at a 3-week interval. The percentages of ( A ) CD3 + T cells, ( B ) CD3 + CD8 + T cells, and ( C ) CD3 + CD4 + T cells among the PBMCs were analyzed by flow cytometry. Data are presented as means and plotted with connecting lines. Each data point represents one sample value. The red triangles represent the time points of inoculation.

Article Snippet: The antibody cocktail included an FITC-conjugated anti-CD3 mAb (CD3-12; Abcam, ab11089) together with a PE-conjugated anti-duck CD4 mAb (Du CD4-2; Bio-Rad, MCA2478) or a PE-conjugated anti-duck CD8 mAb (Du CD8-1; Bio-Rad, MCA2479), which were labeled with a PE/R-phycoerythrin Conjugation Kit (Abcam, ab102918), respectively.

Techniques: Flow Cytometry

Journal: Journal of Virology

Article Title: Recombinant duck enteritis virus harboring the hemagglutinin genes of influenza virus rapidly induces specific cellular immunity in ducks

doi: 10.1128/jvi.02014-25

Figure Lengend Snippet: Specific CD8 + T-cell responses induced by rDEV-dH5/H7 and vDEV. (A) Timeline of inoculation and sample testing. Arrows indicate inoculation, and red circles indicate bleeds, tests, and post-vaccination time points. ( B ) Frequency of specific CD3 + CD8 + T cells expressing IFN-γ + induced by DEV virion. ( C–F ) Frequency of HA-specific CD3 + CD8 + T cells expressing IFN-γ + . PBMCs were stimulated with single inactivated virus antigens: GZ/S4184 (H5N6), LN/SD007 (H5N1), GX/SD098 (H7N9), or a mixture of antigens of these three influenza virus antigens; CD3 + CD8 + IFN-γ + T cells were then quantified. ( G and H ) Uncorrelated control. CD3 + CD8 + IFN-γ + T cells were quantified after PBMCs were stimulated with WSN (H1N1) or La Sota (NDV). Data are presented as means in histograms. Each data point represents one sample value. Statistical significance: * P < 0.05, ** P <0.01, and *** P < 0.001. P values were determined using a two-tailed unpaired Student’s t test. ( I ) Representative gating strategy for HA-specific CD3 + CD8 + T cells expressing IFN-γ + detected at 31 days post-prime vaccination, following stimulation with the influenza virus antigens mixture. RPMI 1640 and PMA plus ionomycin served as negative and positive controls, respectively.

Article Snippet: The antibody cocktail included an FITC-conjugated anti-CD3 mAb (CD3-12; Abcam, ab11089) together with a PE-conjugated anti-duck CD4 mAb (Du CD4-2; Bio-Rad, MCA2478) or a PE-conjugated anti-duck CD8 mAb (Du CD8-1; Bio-Rad, MCA2479), which were labeled with a PE/R-phycoerythrin Conjugation Kit (Abcam, ab102918), respectively.

Techniques: Expressing, Virus, Control, Two Tailed Test

Journal: Journal of Virology

Article Title: Recombinant duck enteritis virus harboring the hemagglutinin genes of influenza virus rapidly induces specific cellular immunity in ducks

doi: 10.1128/jvi.02014-25

Figure Lengend Snippet: Expansion of T cells induced by rDEV-dH5/H7 and vDEV. Four-week-old ducks were inoculated i.m. with two doses of 10 5 TCID 50 of rDEV-dH5/H7 or vDEV at a 3-week interval. The percentages of ( A ) CD3 + T cells, ( B ) CD3 + CD8 + T cells, and ( C ) CD3 + CD4 + T cells among the PBMCs were analyzed by flow cytometry. Data are presented as means and plotted with connecting lines. Each data point represents one sample value. The red triangles represent the time points of inoculation.

Article Snippet: The PBMCs were plated in 1.5 mL centrifuge tubes (10 6 cells/tube) and fixed with 100 μL of 0.3% paraformaldehyde for 20 min at room temperature and subsequently stained for 30 min at room temperature with either mouse anti-duck CD4 mAb (Du CD4-2; Bio-Rad, MCA2478) or mouse anti-duck CD8 mAb (Du CD8-1; Bio-Rad, MCA2479) diluted in PBS containing 0.05% Tween-20.

Techniques: Flow Cytometry

Journal: Journal of Virology

Article Title: Recombinant duck enteritis virus harboring the hemagglutinin genes of influenza virus rapidly induces specific cellular immunity in ducks

doi: 10.1128/jvi.02014-25

Figure Lengend Snippet: Specific CD8 + T-cell responses induced by rDEV-dH5/H7 and vDEV. (A) Timeline of inoculation and sample testing. Arrows indicate inoculation, and red circles indicate bleeds, tests, and post-vaccination time points. ( B ) Frequency of specific CD3 + CD8 + T cells expressing IFN-γ + induced by DEV virion. ( C–F ) Frequency of HA-specific CD3 + CD8 + T cells expressing IFN-γ + . PBMCs were stimulated with single inactivated virus antigens: GZ/S4184 (H5N6), LN/SD007 (H5N1), GX/SD098 (H7N9), or a mixture of antigens of these three influenza virus antigens; CD3 + CD8 + IFN-γ + T cells were then quantified. ( G and H ) Uncorrelated control. CD3 + CD8 + IFN-γ + T cells were quantified after PBMCs were stimulated with WSN (H1N1) or La Sota (NDV). Data are presented as means in histograms. Each data point represents one sample value. Statistical significance: * P < 0.05, ** P <0.01, and *** P < 0.001. P values were determined using a two-tailed unpaired Student’s t test. ( I ) Representative gating strategy for HA-specific CD3 + CD8 + T cells expressing IFN-γ + detected at 31 days post-prime vaccination, following stimulation with the influenza virus antigens mixture. RPMI 1640 and PMA plus ionomycin served as negative and positive controls, respectively.

Article Snippet: The PBMCs were plated in 1.5 mL centrifuge tubes (10 6 cells/tube) and fixed with 100 μL of 0.3% paraformaldehyde for 20 min at room temperature and subsequently stained for 30 min at room temperature with either mouse anti-duck CD4 mAb (Du CD4-2; Bio-Rad, MCA2478) or mouse anti-duck CD8 mAb (Du CD8-1; Bio-Rad, MCA2479) diluted in PBS containing 0.05% Tween-20.

Techniques: Expressing, Virus, Control, Two Tailed Test

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Proportions of CD3+, CD8 +, and CD4 + T cells by flow cytometry in control and aCD40 treated Brpkp110 tumors on day 13 post treatment ( n = 10). B FoxP3 + CD4+ regulatory T cells on day 7 post-treatment ( n = 7). C Proportions of Granzyme B + T cells in subpopulations by flow cytometry in control and aCD40 treated Brpkp110 tumors on day 7 post implantation ( n = 10). D Images of immunofluorescent staining for CD8 (red) and nuclei (blue) and CD8 staining quantification in untreated (control) and aCD40 treated Brpkp110 tumors on day 7 post treatment ( n = 4–6). E Quantification of CD8 immunofluorescent staining in outer, middle, and inner thirds of control and aCD40 treated tumors on day 7 post treatment ( n = 5–6). F After 7 days of treatment with aCD40, Brpkp110 tumors were minced and cultured ex vivo. Supernatant was collected and pooled for each treatment group after 48 h and cytokines were measured ( n = 2, with 3 tumors pooled per group). G T cell activation and proliferation markers measured by flow cytometry in CD4+ and CD8 + T cell populations of control and aCD40 treated Brpkp110 TDLN on day 7 post-treatment ( n = 10). H Growth curves of control and aCD40 treated Brpkp110 tumors implanted into WT hosts with or without T cell depletions ( n = 17–20). I Tumor growth curve of aCD40 (treatment on Day 8) +/− FTY720 (treatment started on Day 7) treated Brpkp110 tumors ( n = 14–16). Data: ( A – C , D (right), E – G ) median, ( H , I ) mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Flow Cytometry, Control, Staining, Cell Culture, Ex Vivo, Activation Assay

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 26 post implantation (right) of Brpkp110 tumors. Indicated treatments initiated on day 7 post implantation ( n = 18–20). B Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 26 post implantation (right) of E0771 tumors. Indicated treatments initiated on day 8 post implantation ( n = 12–15). C Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 31 post implantation (right) of AT3 tumors. Indicated treatments initiated on day 9 post implantation ( n = 14). D Tumor growth curves (left) and tumor volume changes compared to pretreatment on day 34 post implantation (right) of EpH4 1424 tumors. Indicated treatments initiated on day 5 post implantation ( n = 14–16). E Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 27 post implantation in control and aCD40+ICB treated hosts with and without CD8 + T cell depletions. Indicated treatments initiated on day 7 post implantation ( n = 12–15). F Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 25 post implantation in control and aCD40+ICB treated hosts with and without CD4 + T cell depletions. Indicated treatments initiated on day 8 post implantation ( n = 14–18). G Brpkp110 tumor growth curves (left) and volume changes compared to pretreatment (right) on day 27 post implantation in control and aCD40+ICB treated hosts with and without CD4+ and CD8 + T cell depletions. Indicated treatments initiated on day 7 post implantation ( n = 12–18). H Tumor growth curves (left) and volume changes compared to pretreatment (right) on day 26 post Brpkp110 tumor implantation into WT and BATF3 KO hosts. Indicated treatments initiated on day 7 post implantation ( n = 14–18). Data: ( A – H left) mean ± SEM, ( A – H right) each column represents individual tumor. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Control, Tumor Implantation

Journal: NPJ Breast Cancer

Article Title: Agonistic CD40 elicits CD8+ T-cell-dependent primary responses and CD4+ T-cell-dependent long-term immunity in breast cancer

doi: 10.1038/s41523-025-00889-7

Figure Lengend Snippet: A Proportions of circulating effector memory (CD44 + CD62L-), central memory (CD44 + CD62L+), and naïve (CD44-CD62L-) CD4+ (left) and CD8+ (right) in blood, 3 months post treatment induced tumor clearance ( n = 5–6, data representative of 2 experiments with similar results). B Secondary Brpkp110 tumor rechallenge of naïve and previously Brpkp110 tumor-bearing mice cured after aCD40 + ICB, at least 2 months post primary tumor clearance ( n = 12–14, data representative of 3 experiments with similar results). C Control and rechallenge tumor growth in T cell sufficient ( n = 6–12) and T cell depleted hosts ( n = 12–14, data representative of 2 experiments with similar results). D Brpkp110 tumor growth curves in intra-tumoral (IT) vehicle (control) and IT aCD40 treated hosts. aCD40 administered tumors denoted as aCD40 IT and contralateral untreated tumors denoted as CD40 IT Distant ( n = 9–12, data representative of 2 experiments with similar results). E Brpkp110 tumor growth curves in intra-tumoral (IT) vehicle (control) and IT or intraperitoneal (IP) aCD40 or ICB received hosts ( n = 4–9, data representative of 2 experiments with similar results). Data: ( A ) median, ( C – E ) mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001.

Article Snippet: Tissue sections were blocked with 5% donkey serum diluted in PBS with 0.3% Triton X-100 and stained with rabbit anti-mouse primary CD8 (clone: D4W2Z, Cell Signaling Technology 98941, 1:100) followed by donkey anti-rabbit Alexa Fluor 594 (Thermo Scientific A-21207, 1:250) secondary antibody.

Techniques: Control

Journal: Journal for Immunotherapy of Cancer

Article Title: Targeting RRBP1 reverses immune evasion and enhances immunotherapy efficacy via the CXCL10-CXCR3 axis in bladder cancer

doi: 10.1136/jitc-2025-013809

Figure Lengend Snippet: Tumor-intrinsic RRBP1 inhibition triggers antitumor immunity. ( A ) Representative images of IHC staining for RRBP1 and CD8 + T cells in BC samples. ( B ) The correlation between RRBP1 expression and CD8 + T-cell infiltration was analyzed based on 96 patients from in-house BC cohort. Scale bar: 50 µm. ( C ) Representative images of IHC staining for RRBP1 expression in PD, SD, PR, and CR samples. Scale bar: 50 µm. ( D ) Bar plot showed the response rates of anti-PD-L1 therapy. Blue bars represent CR/PR, Red bars represent PD/SD. ( E ) Volcano plot of RNA-seq data for shNC or shRRBP1 tumors (n=3). Differentially expressed genes were identified with the threshold of |log2 (fold change) | >1 and FDR<0.05. ( F ) GSEA for DEGs showed the activation of immune-associated pathways in shRRBP1 tumors in the RNA-seq data. ( G ) Representative images of IHC and mIHC staining for RRBP1 and CD8 + T cells in shNC, shRRBP1, control or radezolid tumor tissues. Expression levels of the indicated proteins were displayed. Scale bar: 20 µm. ( H, I ) Flow cytometry showed the percentages of CD8 + T cells in CD3 + cells in shNC, shRRBP1, control or radezolid tumor tissues. Data are represented as mean means±SD. Statistical analysis was performed using Spearman correlation analysis ( B ), unpaired two-tailed t-test ( I ). ****p<0.0001. BC, bladder cancer; CR, complete response; FDR, false discovery rate; progressive disease; PR, partial response; PD-L1, programmed death-ligand 1; RNA-seq, RNA sequencing; RRB1, ribosomal-binding protein 1; SD, stable disease; IHC, immunohistochemistry; GSEA, gene set enrichment analysis; DEGs, differentially expressed genes; mIHC, multiplex immunohistochemistry.

Article Snippet: CD8 + T-cell depletion was achieved by intraperitoneal injection of anti-mouse CD8 mAbs (200 μg per mouse; Bio X Cell, catalog no. BE0117, clone: YTS169.4) administered on days –6, –3, and –1 prior to tumor inoculation, with additional injections on days 7, 9, and 11 post-inoculation.

Techniques: Inhibition, Immunohistochemistry, Expressing, RNA Sequencing, Activation Assay, Staining, Control, Flow Cytometry, Two Tailed Test, Binding Assay, Multiplex Assay

Journal: Journal for Immunotherapy of Cancer

Article Title: Targeting RRBP1 reverses immune evasion and enhances immunotherapy efficacy via the CXCL10-CXCR3 axis in bladder cancer

doi: 10.1136/jitc-2025-013809

Figure Lengend Snippet: Single-cell RNA sequencing reveals the difference of CD8 + T-cell subgroup. The UMAP plot of CD8 + T cells subpopulation, color-coded by cell cluster and cell type. ( A ) The expression of markers in each CD8 + T cells subpopulation. ( B ) Bar plot showed the proportion of CD8 + T cells subpopulation in the shNC and shRRBP1 groups. ( C ) The percentage of each CD8 + T-cell clusters in shNC and shRRBP1 groups. ( D ) Heatmap showed the differentially activated pathway among all the CD8 + T-cell clusters. ( E ) The differentially expressed genes in CD8 + T cells between shNC and shRRBP1 groups. ( F ) KEGG analysis for differentially expressed genes showed the enrichment of immune-associated pathways. ( G, H ) mIHC and flow cytometric analysis displayed the tumor-infiltrating IFN-γ + or GZMB + CD8 + T cells in shNC or shRRBP1 tumor tissues. Scale bar: 20 µm. ( I–K ) C57BL/6 mice were subcutaneously injected with 5×10 5 stable MB49 cells (shNC or shRRBP1 cells) (n=6). Isotype control (IgG) or anti-mouse CD8 antibody administered on days –6, –3, and –1 before tumor challenge, with the same dose repeated on days 7, 9 and 11 after tumor challenge. Tumor sizes ( I ), volumes ( J ), and weight ( K ) were measured. Data are represented as mean means±SD. Statistical analysis was performed using unpaired two-tailed t-test ( H, K ) and two-way ANOVA with Tukey’s multiple comparison test ( J ). *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. ANOVA, analysis of variance; GZMB, Granzyme B; IFN, interferon; TEX, exhausted T cells; UMAP, Uniform Manifold Approximation and Projection; mIHC, multiplex immunohistochemistry; KEGG, Kyoto Encyclopedia of Genes and Genomes.

Article Snippet: CD8 + T-cell depletion was achieved by intraperitoneal injection of anti-mouse CD8 mAbs (200 μg per mouse; Bio X Cell, catalog no. BE0117, clone: YTS169.4) administered on days –6, –3, and –1 prior to tumor inoculation, with additional injections on days 7, 9, and 11 post-inoculation.

Techniques: Single Cell, RNA Sequencing, Expressing, Injection, Control, Two Tailed Test, Comparison, Multiplex Assay, Immunohistochemistry

Journal: Journal for Immunotherapy of Cancer

Article Title: Targeting RRBP1 reverses immune evasion and enhances immunotherapy efficacy via the CXCL10-CXCR3 axis in bladder cancer

doi: 10.1136/jitc-2025-013809

Figure Lengend Snippet: RRBP1 inhibition promotes antitumor immunity via the CXCL10-CXCR3 axis in BC. ( A ) ScRNA-seq data showed the CXCR3 expression of CD8+T cells in shNC and shRRBP1 groups. ( B ) The correlation between CXCR3 expression and CXCL10 expression or activated CD8 + T cell based on 571 patients from TCGA-BLCA cohort and GSE13507 cohorts. ( C ) MB49 cells were co-cultured with CD8 + T cells, and tumor cells were stained with crystal violet. ( D ) Evaluation of the effect of genetic inhibition of RRBP1 on the cytotoxicity of CD8 + T cells in vitro conditioned culture model. ( E ) Schematic diagram of in vitro CD8 + T-cell migration assays. ( F ) The number of CD8 + T cells passing through the membrane of a Transwell system was analyzed by flow cytometry. ( G–I ) C57BL/6 mice were subcutaneously injected with 5×10 5 stable MB49 cells (shNC or shRRBP1 cells) (n=6). Tumor-bearing mice received intraperitoneal injection of either vehicle or anti-CXCL10 when the tumor volume reached a calculated average of 100 mm 3 . The tumor sizes ( G ), volumes ( H ), and weights ( I ) were measured. ( J ) Representative images of IHC and mIHC staining for CD8, CXCR3, CXCL10, IFN-γ, GZMB in different tumor tissues. ( K ) Flow cytometric analysis of tumor-infiltrating CD8 + T cells, CXCR3 + CD8 + T cells, IFN-γ + CD8 + T cells or GZMB + CD8 + T cells in distinct tumor tissues. Data are represented as mean means±SD. Statistical analysis was performed using unpaired two-tailed t-test ( D, F, I, K ) and two-way ANOVA with Tukey’s multiple comparison test ( H ). The data presented represent on one or three independent experiments. *p<0.01, **p<0.01, ***p<0.001. ANOVA, analysis of variance; BC, bladder cancer; GZMB, Granzyme B; IFN, interferon; RRBP1, ribosomal-binding protein 1; scRNA-seq, single-cell RNA sequencing; IHC, immunohistochemistry; mIHC, multiplex immunohistochemistry; BLCA, bladder urothelial carcinoma.

Article Snippet: CD8 + T-cell depletion was achieved by intraperitoneal injection of anti-mouse CD8 mAbs (200 μg per mouse; Bio X Cell, catalog no. BE0117, clone: YTS169.4) administered on days –6, –3, and –1 prior to tumor inoculation, with additional injections on days 7, 9, and 11 post-inoculation.

Techniques: Inhibition, Expressing, Cell Culture, Staining, In Vitro, Migration, Membrane, Flow Cytometry, Injection, Two Tailed Test, Comparison, Binding Assay, Single Cell, RNA Sequencing, Immunohistochemistry, Multiplex Assay

Journal: Journal for Immunotherapy of Cancer

Article Title: Targeting RRBP1 reverses immune evasion and enhances immunotherapy efficacy via the CXCL10-CXCR3 axis in bladder cancer

doi: 10.1136/jitc-2025-013809

Figure Lengend Snippet: RRBP1 inhibition enhances response to anti-PD-L1 therapy in BC. ( A–D ) The protein expression of surface PD-L1 was analyzed in BC cells or tumor tissues by flow cytometry after RRBP1 inhibition and was shown as the mean fluorescence intensity. ( E–G ) C57BL/6 mice were subcutaneously injected with 5×10 5 stable MB49 cells (shNC or shRRBP1 cells) (n=6). Tumor-bearing mice were received intraperitoneal injection of either vehicle or anti-PD-L1 antibody when the tumor volume reached a calculated average of 100 mm 3 . The tumor sizes ( E ), volumes ( F ), and weights ( G ) were measured. ( H ) Representative images of IHC and mIHC staining for CD8, CXCR3, CXCL10, IFN-γ, GZMB in different tumor tissues. ( I ) Flow cytometric analysis of tumor-infiltrating CD8 + T cells, CXCR3 + CD8 + T cells, IFN-γ + CD8 + T cells or GZMB + CD8 + T cells in distinct tumor tissues. Data are represented as mean means±SD. Statistical analysis was performed using unpaired two-tailed t-test ( B, D, G, I ) and two-way ANOVA with Tukey’s multiple comparison test ( F ). The data presented represent on one or three independent experiments. *p<0.01, **p<0.01, ***p<0.001. ANOVA, analysis of variance; BC, bladder cancer; GZMB, Granzyme B; IFN, interferon; PD-L1, programmed death-ligand 1; RRBP1, ribosomal-binding protein 1; IHC, immunohistochemistry; mIHC, multiplex immunohistochemistry.

Article Snippet: CD8 + T-cell depletion was achieved by intraperitoneal injection of anti-mouse CD8 mAbs (200 μg per mouse; Bio X Cell, catalog no. BE0117, clone: YTS169.4) administered on days –6, –3, and –1 prior to tumor inoculation, with additional injections on days 7, 9, and 11 post-inoculation.

Techniques: Inhibition, Expressing, Flow Cytometry, Fluorescence, Injection, Staining, Two Tailed Test, Comparison, Binding Assay, Immunohistochemistry, Multiplex Assay