Journal: Cancer Immunology, Immunotherapy : CII

Article Title: The role of Ly49E receptor expression on murine intraepithelial lymphocytes in intestinal cancer development and progression

doi: 10.1007/s00262-016-1894-6

Figure Lengend Snippet: Tumor-infiltrating T cells and tumor uPA expression in AOM-treated Ly49E WT versus Ly49E KO mice. a Hematoxylin/eosin-(H&E)- (upper), and CD3-(lower) stained paraffin tumor sections from AOM-treated Ly49E WT and Ly49E KO mice. Scale bar 250 µm, ×100 magnification. A graph showing CD3 mean gray value/mm2 according to tumor surface area (mm2) is shown for AOM-treated Ly49E WT and Ly49E KO mice (n = 6). b Colon tumor-infiltrating IEL subpopulation frequencies in AOM-treated Ly49E WT and Ly49E KO mice 14–22 weeks following the start of treatment, and colon IEL subpopulation frequencies from untreated Ly49E WT and Ly49E KO mice (mean ± SEM; n = 5 for AOM-treated mice; n = 3 for untreated mice). The percentage of TCRαβ and TCRγδ IEL is shown as a fraction of the total numbers of T cells. TCRαβ CD4, TCRαβ CD8αβ and TCRαβ CD8αα IEL subpopulation frequencies are shown as a percentage of the total TCRαβ IELs. TCRγδ DN and TCRγδ CD8αα IEL subpopulation frequencies are shown as a percentage of the total TCRγδ IEL. c Dot plots are shown for CD4/CD8β versus CD8α expression in colon tumor-infiltrating IEL in AOM-treated Ly49E WT and Ly49E KO mice, and colon IEL from untreated Ly49E WT and Ly49E KO mice. Numbers indicate the percentage of cells in each quadrant. Dot plots are representative for n = 5 AOM-treated mice and n = 3 untreated mice. d Tumor uPA expression in tumors of varying size from AOM-treated Ly49E WT and Ly49E KO mice at 14–22 weeks following the start of treatment. n.s. not significant. Data were analyzed using the nonparametric two-tailed Mann–Whitney U-test or the Kruskall–Wallis test. A p value ≤0.05, (*), a p value ≤0.01 (**) and p value ≤0.001 (***), were considered statistically significant

Article Snippet: Anti-CD8α (PE/Cy7-conjugated, clone 53-6.7) from eBioscience (San Diego, CA, USA).

Techniques: Expressing, Staining, Two Tailed Test, MANN-WHITNEY

Journal: PLoS ONE

Article Title: Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway

doi: 10.1371/journal.pone.0273075

Figure Lengend Snippet: Antibodies used for flow cytometry.

Article Snippet: Pooled DCs were washed in flow buffer (PBS with 0.5% FBS), incubated with anti-mouse Fc Block (Thermo Fisher Scientific), and then stained with CD8α PE-Cy7 (Thermo Fisher; clone: 53–6.7), and CD24 Pacific Blue (Biolegend; clone: M1/69).

Techniques: Cytometry

Journal: PLoS ONE

Article Title: Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway

doi: 10.1371/journal.pone.0273075

Figure Lengend Snippet: Splenic pan DCs were isolated from naïve BALB/c mice and analyzed by flow cytometry to characterize pre-cDC1s and CD8α+ cDC1s. (A) Representative flow plot demonstrating gating strategy. Gated on CD11c+B220- (conventional DCs); pre-cDC1s are gated as CD24 high CD8α- and CD8α+ cDC1s are gated as CD8α+. (B) Mean absolute cell number of respective DC subsets shown with SEM. (C-F) Mean percent and MFI with representative histograms shown with SEM for expression of (C) PD-L1, (D) PIR-B, (E) CD70, and (F) ICOSL on CD8α+ cDC1s (black) and pre-cDC1s (cyan). Data is representative of two experiments. (n = 5 per group). Paired t tests were used to determine significance between DC subsets. **P<0.01, ***P<0.001.

Article Snippet: Pooled DCs were washed in flow buffer (PBS with 0.5% FBS), incubated with anti-mouse Fc Block (Thermo Fisher Scientific), and then stained with CD8α PE-Cy7 (Thermo Fisher; clone: 53–6.7), and CD24 Pacific Blue (Biolegend; clone: M1/69).

Techniques: Isolation, Flow Cytometry, Expressing

Journal: PLoS ONE

Article Title: Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway

doi: 10.1371/journal.pone.0273075

Figure Lengend Snippet: BALB/c mice were dosed with 200 μg of 2.43 mAb antibody intraperitoneally on day -3 and day -1. On day 0, splenic DCs were isolated and analyzed by flow cytometry. Data is representative of three independent experiments. (n = 4 per group) (A) Representative flow plots depicting relative cDC1 proportions in naïve DCs (left) and 2.43 mAb DCs (right). Gated on CD11c+B220- (conventional DCs). (B) Mean percent of CD8α+ cDC1s (black) and pre-cDC1s (cyan) in untreated and 2.43 mAb-treated DCs shown with SEM. Two-way ANOVA and Dunnett’s multiple comparisons used to determine significance. **P<0.01, ****P<0.0001. (C) Mean absolute cell number of pre-cDC1s (CD24 high CD8α-) in untreated (black) and 2.43 mAb-treated (magenta) groups shown with SEM. Unpaired t test used to determine significance. *P<0.05. (D) Schematic depicting experimental design for proposed allogeneic mixed leukocyte reaction (MLR) using 2.43 mAb treatment.

Article Snippet: Pooled DCs were washed in flow buffer (PBS with 0.5% FBS), incubated with anti-mouse Fc Block (Thermo Fisher Scientific), and then stained with CD8α PE-Cy7 (Thermo Fisher; clone: 53–6.7), and CD24 Pacific Blue (Biolegend; clone: M1/69).

Techniques: Isolation, Flow Cytometry

Journal: PLoS ONE

Article Title: Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway

doi: 10.1371/journal.pone.0273075

Figure Lengend Snippet: BALB/c mice were dosed with 200 μg of 2.43 mAb antibody intraperitoneally on day -3 and day -1. On day 0, splenic DCs were isolated and plated in an allogeneic MLR. DC composition was determined by flow cytometry on day 2 of co-culture. Data is representative of two independent experiments. (n = 4 per group) (A) Representative flow plots depicting percent of H2K d +CD11c+ DCs in naïve DCs (left) and 2.43 mAb DCs (right) on day 2 of co-culture. (B) Mean percent of H2K d +CD11c+ DCs for naïve (black) or 2.43 mAb DCs (magenta) shown with SEM. Unpaired t test was used to determine significance between groups. **P<0.01. (C) Representative flow plots depicting percent CD8α+ cDC1s and pre-cDC1s in naïve (left) and 2.43 mAb DCs (right). (D) Mean percent CD8α+ cDC1s (black bars) and pre-cDC1s (teal bars) in naïve and 2.43 mAb DCs shown with SEM. 2way ANOVA and Šidák’s multiple comparison used to determine statistical significance. **P<0.01.

Article Snippet: Pooled DCs were washed in flow buffer (PBS with 0.5% FBS), incubated with anti-mouse Fc Block (Thermo Fisher Scientific), and then stained with CD8α PE-Cy7 (Thermo Fisher; clone: 53–6.7), and CD24 Pacific Blue (Biolegend; clone: M1/69).

Techniques: Isolation, Flow Cytometry, Co-Culture Assay

Journal: PLoS ONE

Article Title: Murine precursors to type 1 conventional dendritic cells induce tumor cytotoxicity and exhibit activated PD-1/PD-L1 pathway

doi: 10.1371/journal.pone.0273075

Figure Lengend Snippet: Splenic DCs were isolated from naïve BALB/c mice and pre-cDC1s and CD8α+ cDC1s were cell sorted for RNA sequencing. (A) Enhanced volcano plot identifying genes that are up or down-regulated in pre-cDC1s compared to CD8α+ cDC1s. (B-C) Pathway results using Qiagen’s IPA software. P = 0.05 corresponds to a 1.3 -log p-value, and anything above this value is considered statistically significant. (B) Pathways predicted to be significantly inhibited (z-score < -2) in pre-cDC1s compared to CD8α+ cDC1s. (C) Pathways predicted to be significantly activated (z-score < 2) in pre-cDC1s compared to CD8α+ cDC1s. (D) Heat map showing relative gene expression levels for the PD-1/PD-L1 cancer immunotherapy pathway. Full documentation, methodology, and raw data for this data set is available online at: https://doi.org/10.25422/azu.data.14241902 .

Article Snippet: Pooled DCs were washed in flow buffer (PBS with 0.5% FBS), incubated with anti-mouse Fc Block (Thermo Fisher Scientific), and then stained with CD8α PE-Cy7 (Thermo Fisher; clone: 53–6.7), and CD24 Pacific Blue (Biolegend; clone: M1/69).

Techniques: Isolation, RNA Sequencing Assay, Software, Expressing