Journal: Cell

Article Title: Transcriptional Basis of Mouse and Human Dendritic Cell Heterogeneity

doi: 10.1016/j.cell.2019.09.035

Figure Lengend Snippet: Single-Cell Survey Reveals Heterogeneity of cDC2s with Two Subsets Delineated by Expression of T-Bet (A) Representative contour plot showing gating strategy for splenic DCs in Tbx21 RFP-Cre mice. DCs defined as Lin(CD3,CD19,CD49b,Siglec-F) – Ly6C – CD64 – CD11c + MHCII + . (B) Frequency of T-bet + cDC2s across tissues. Each circle represents one mouse. In the peripheral and mesenteric LN (PLN and MLN), migratory DCs were defined as MHCII hi CD11c int and resident DCs as MHCII int CD11c hi . Error bars represent mean ± SEM. (C) Analysis of RFP + and YFP + splenic cDC2s from Tbx21 RFP-CreERT2 Rosa26 YFP mice, 3 days post tamoxifen gavage. (D) Percent RFP + and YFP + of cDC2 cells. Percent RFP + of YFP + cDC2s at indicated time points post tamoxifen gavage (right). Error bars represent mean ± SEM; n = 3–4 mice per time point. (E) t-SNE embedding of 4,464 DCs. Colors indicate unsupervised clustering by Phenograph (left panel) or classification based on expression of canonical markers (right panel). (F) Expression of canonical DC markers across the transcriptionally defined DC clusters from (E). (G) Proportion of T-bet (RFP + ) cells in each cell cluster identified in (D). (H) Violin plot showing expression of the cell-cycle signature across the DC clusters from (E). (I) Similarity of bulk T-bet – cDC2s, T-bet + cDC2, and cDC1 transcriptomes to the reference single-cell DC clusters (E). Colors represent the correlation coefficient between the cell population identified in the row label and the DC cluster identified by the column label. See also and .

Article Snippet: Anti-mouse CD3ε (PE-Cy7) , Tonbo Biosciences , Cat#60-0031; RRID: AB_2621824 ; Clone 145-2C11.

Techniques: Expressing

Journal: Cell

Article Title: Transcriptional Basis of Mouse and Human Dendritic Cell Heterogeneity

doi: 10.1016/j.cell.2019.09.035

Figure Lengend Snippet: Single-Cell Survey Reveals Heterogeneity of cDC2s, Related to Figure 1 A. Representative histogram showing expression of T-bet (RFP) in splenic cells from Tbx21 RFP-cre mice. (B). Expression of T-bet in CD11b + XCR1 + DCs from the intestinal lamina propria. Data representative of > 5 independent experiments, with at least 3 mice per experiment. (C). Expression of T-bet in splenic myeloid cells. Cells were defined as: (i) Ly-6C hi monocytes (Lin – Ly6C + Ly6G – CD11b + CX3CR1 + ); neutrophils (Lin – Ly6C + Ly6G + ); macrophages (Lin – CD64 + Ly6C – ). Lineages (Lin) were defined as: CD3e, CD90.2, CD19, CD49b and Siglec F. Each circle represents an individual mouse, error bars represent mean ± SEM. (D). Left: Gating strategy for single-cell sorting. DCs were defined as Lin(CD3, CD19, CD90) – Ly6C – CD64 – CD11c + MHCII + . Two populations were sampled: RFP + DCs and RFP – DCs (encompassing XCR1 + cDC1s, CD11b + RFP – and CD11b – XCR1 – DCs). Right: Post-sort purity of RFP + and RFP – cells. Contaminating population of Ly6C + cells identifiable on post-sort purity (lower panel). (E). Similarity of splenic CD11c + MHCII + cells to reference myeloid cells (ImmGen Consortium) Colors represent the Pearson correlation between the mean gene expression from the dendritic cell cluster in the rows and the bulk reference transcriptome in the columns. (F). Top 20 positive and negative gene loadings of PC1 for T-bet + cDC2 clusters after cell-cycle correction (left panel). Scatterplot of PC1 and PC2 for T-bet + cDC2 clusters after cell-cycle correction (right panel).

Article Snippet: Anti-mouse CD3ε (PE-Cy7) , Tonbo Biosciences , Cat#60-0031; RRID: AB_2621824 ; Clone 145-2C11.

Techniques: Expressing, FACS, Gene Expression

Journal: Cell

Article Title: Transcriptional Basis of Mouse and Human Dendritic Cell Heterogeneity

doi: 10.1016/j.cell.2019.09.035

Figure Lengend Snippet: Environmental Cues Drive Distinct DC2 Differentiation Pathways within the Spleen, Related to Figure 5 (A). Gating strategy for the identification of DC progenitors in the bone marrow (BM) (B). Palantir pseudo-time analysis of differentiation potential and branch probabilities from the Siglec-H + pre-DC state to T-bet + cDC2 and T-bet – cDC2 terminal states. (C). Plots showing Palantir differentiation potential (y axis) along Palantir pseudo-time (x axis) for Siglec-H + DC and T-bet + cDC2s (top) or Siglec-H + DC and T-bet – cDC2 clusters (bottom) (D). Plots showing the top two diffusion component embeddings for Siglec-H + DC and T-bet + cDC2 clusters (top) or Siglec-H + DC and Tbet – cDC2 clusters (bottom). Black arrow indicates Siglec-H + DC cluster cells adjacent to cells from the proliferative T-bet + cDC2 clusters 6 and 8. (E). Top panel: plots showing probability of each cell being within 20 nearest neighbors of randomly sampled shortest paths from the Siglec-H + DC to the indicated end points. Middle panel: plots showing the proportion of cells belonging to Siglec-H + DC, T-bet + cDC2, or T-bet – cDC2 from 20 nearest neighbors of randomly sampled shortest paths. Bottom: plots showing diffusion distance step sizes for each step along the indicated shortest paths (bottom panel). Colors illustrate cluster membership. (F). Graph showing AUC (x axis) for genes differentially expressed between Siglec-H + DC cluster (cluster 11) and all other cDC2 clusters. EMD on the y axis. Dashed lines represents μ EMD ± 3σ EMD . (G). Gating strategy for FACS-isolation of MHCII + ILC3s: Lin = CD3, CD19, CD49b, Siglec-F. (H). Heatmap reports scaled expression of 3550 differentially expressed genes (log 2 FC > 1, FDR < 0.01) between ILC3s and Rorγt fm cDC2s. Selected genes listed to the right. (I). Representative flow cytometric analysis of phenotypes of splenic progeny from Tbx21 RFP-cre CD45.2 + Ly6C − CD64 – MHCII + CD11c + Siglec-H + pre-DCs adoptively transferred into sub-lethally irradiated CD45.1 recipient mice 7 days earlier (data from one experiment with n = 3). J. Sort purified T-bet + or T-bet – cDC2 were cultured for 24hrs in the presence of LPS, CpG, TNF-α or IFN−γ. Representative overlay histogram showing the expression of RFP(T-bet) at 24hrs. Data representative of 2 (TNF-α) or 4 (all other cytokines/TLR agonists) independent experiments, n = 2-3.

Article Snippet: Anti-mouse CD3ε (PE-Cy7) , Tonbo Biosciences , Cat#60-0031; RRID: AB_2621824 ; Clone 145-2C11.

Techniques: Diffusion-based Assay, Isolation, Expressing, Irradiation, Purification, Cell Culture

Journal: Cell

Article Title: Transcriptional Basis of Mouse and Human Dendritic Cell Heterogeneity

doi: 10.1016/j.cell.2019.09.035

Figure Lengend Snippet: Human DC Heterogeneity, Related to Figure 7 (A). Violin plots showing expression distribution of mouse DC subset marker genes across human peripheral blood DC and monocyte clusters identified in Villani et al. (2017) . (B). Representative flow cytometric analysis of mouse peripheral blood cDC2s showing absence of T-bet (RFP) + cDC2s. (C). Gating strategy for FACS-isolation of human spleen DCs for scRNA-seq. DCs were defined as live, LIN(CD3,CD56,CD19) − CD14 – CD11C + HLA-DR + . (D). Representative flow cytometry analysis of human spleen cDC2s gated as Lin(CD3,CD56,CD19) – CD14 – CD11c + HLA-DR + CD123 – XCR1 – CLEC4A + cells. Left panel: cell surface expression of CD1c and CLEC10A by cDC2s. Right panel: overlay of CLEC10A + and CLEC10A – cDC2s distinguished by differential expression of CLEC4A and FcεR1a. Summary bar graphs show frequency of CD1C + CLEC10A + and CD1C + CLEC10A – cDC2s as a percentage of cDC2s (n = 4 individuals). (E). t -SNE embedding of 9,315 FACS-isolated CD45 + immune cells from two melanoma tumors. Colors indicate unsupervised clustering by Phenograph (left panel) or classification based on expression of canonical markers and correlations with bulk RNA-seq data (right panel). Each dot represents an individual cell. (F). Pearson correlations between cluster centroids in (F) and bulk RNA-seq data from purified immune populations ( Jeffrey et al., 2006 , Novershtern et al., 2011 ) (G). t-SNE map of 2,122 myeloid cells identified in (F). Colors indicate patient sample (left) or unsupervised clustering by Phenograph (right panel). Each dot represents an individual cell. (H). Heatmap of normalized, log transformed and MAGIC imputed expression of top 20 differentially expressed genes, defined by the highest earth mover’s distance (EMD), per Phenograph cluster in E. The colored bar at the top of the heatmap shows assignment of cells to clusters labeled in F, right panel. (I). t-SNE map of human melanoma myeloid cells (H) colored by imputed expression of labeled genes.

Article Snippet: Anti-mouse CD3ε (PE-Cy7) , Tonbo Biosciences , Cat#60-0031; RRID: AB_2621824 ; Clone 145-2C11.

Techniques: Expressing, Marker, Isolation, Flow Cytometry, Quantitative Proteomics, RNA Sequencing, Purification, Transformation Assay, Labeling

Journal: Cell

Article Title: Transcriptional Basis of Mouse and Human Dendritic Cell Heterogeneity

doi: 10.1016/j.cell.2019.09.035

Figure Lengend Snippet:

Article Snippet: Anti-mouse CD3ε (PE-Cy7) , Tonbo Biosciences , Cat#60-0031; RRID: AB_2621824 ; Clone 145-2C11.

Techniques: Recombinant, Staining, Multiplex Assay, Cell Isolation, Gene Expression, Software

Journal: bioRxiv

Article Title: Porcine intestinal innate lymphoid cells and lymphocyte spatial context revealed through single-cell RNA sequencing

doi: 10.1101/2022.01.09.475571

Figure Lengend Snippet: (A) Cell compositions of scRNA-seq data from whole ileum (top), PP (middle), and non-PP (bottom) samples. Cells from each sample type (depicted on the far left) were combined from a total of two animals and overlaid onto t-SNE coordinates originally presented in . The total numbers of cells derived from the total of two animals for each sample type are listed on the far left. On the t-SNE plots, each point represents a single cell; the color of each point corresponds to cell lineage (left t-SNE), cell type (center t-SNE), or cell density (right t-SNE). (B) Pie charts showing proportions of cells from each annotated cell lineage within total cells derived from each sample type in A . The color of a pie slice indicates cell lineage. The total area of each pie chart is not proportional to the total number of cells derived from each sample type. Proportions were calculated from total cells derived from two pigs for each sample type. (C) Plot of the percentage of B cells (CD79α + ) within total leukocytes (CD45 + ; y-axis) from samples of whole ileum, PP, and non-PP (x-axis), as assessed by flow cytometry gating shown in Supplement to Figure 4 - Figure 2A . Measurements from different sample types derived from the same animal are connected by a light grey line. (D) IHC staining for B cell CD79α protein (brown) in a region of ileum with Peyer’s patches (left) or without Peyer’s patches (right). (E) Plot of the percentage of T cells (CD3ε + ) within total leukocytes (CD45 + ; y-axis) from samples of whole ileum, PP, and non-PP (x-axis), as assessed by flow cytometry gating shown in Supplement to Figure 4-Figure 2A . Measurements from different sample types derived from the same animal are connected by a light grey line. (F) IHC staining for T cell CD3ε protein (brown) in a region of ileum with Peyer’s patches (left) or without Peyer’s patches (right). (G) Plot of the percentage of CD4 αβ T cells (left), CD8 αβ T cells (center), or γδ T cells (right) within total T cells (y-axis) of the porcine ileum scRNA-seq dataset. Percentages from samples of whole ileum, PP, and non-PP are shown on the x-axis. CD4 αβ T cells included cells annotated as follicular CD4 αβ T cells, activated CD4 αβ T cells, or cycling CD4 αβ T cells and cells annotated as naïve CD4/CD8 αβ T cells with prediction probability to porcine PBMC CD4 + αβ T cells > prediction probability to porcine PBMC CD8αβ + αβ T cells. CD8 αβ T cells included cells annotated as activated CD8 αβ T cells, cytotoxic CD8 αβ T cells, or cycling CD8 αβ T cells and cells annotated as naïve CD4/CD8 αβ T cells with prediction probability to porcine PBMC CD8αβ + αβ T cells > prediction probability to porcine PBMC CD4 + αβ T cells. γδ T cells included cells annotated as activated γδ T cells, cytotoxic γδ T cells, cycling γδ T cells, SELL hi γδ T cells, and CD2 - γδ T cells. Measurements from different sample types derived from the same animal are connected by a light grey line. (H) Plot of the percentage of CD4 αβ T cells (γδTCR - CD4 + ; left), CD8 αβ T cells (γδTCR - CD8β + ; center), or γδ T cells (γδTCR + ; right) within total T cells (CD3ε + ; y-axis) from samples of whole ileum, PP, and non-PP (x-axis), as assessed by flow cytometry gating shown in Supplement to Figure 4-Figure 2B . Measurements from different sample types derived from the same animal are connected by a light grey line. (I) RNA ISH staining for TRDC (top, red), CD8B (bottom, green), or CD4 (bottom, red) transcripts in regions of ileum with Peyer’s patches (left) or regions of ileum without Peyer’s patches (right). (J) Differential abundance analysis of cell types from porcine ileum scRNA-seq PP versus non-PP samples. Annotated cell types are listed on the y-axis. Each point represents an individual cell neighborhood, where a neighborhood was assigned as a specific cell type if >70% of cells within the neighborhood belonged to the specified cell type annotation. Cell neighborhoods with <70% of cells belonging to a single cell type are not shown. Grey points indicate cell neighborhoods that were not significantly more abundant in a specific sample type. Non-grey points indicate cell neighborhoods exhibiting differential abundance (p<0.1), and red/blue fill of differentially abundant points corresponds to the magnitude and direction of logFC (also corresponding to values listed on the x-axis). Red indicates increased abundance in PP samples, while blue indicates increased abundance in non-PP samples. On the far right, counts of cell neighborhoods with increased abundance in PP samples/no differential abundance/increased abundance in non-PP samples are shown for each cell type. Cycling γδ T cells and cycling group 1 ILCs are not shown on the y-axis due to no cell neighborhoods being assigned to these cell types. scRNA-seq data shown in A-B, G, & J were derived from ileum of two seven-week-old pigs. Images shown in I were also taken from a seven-week-old pig used for ileum scRNA-seq. Flow cytometry and IHC experiments were not performed on animals used for scRNA-seq. Flow cytometry experiments shown in C & E were conducted using four six-week-old pigs. Flow cytometry data shown in H was performed using five nine-week-old pigs. IHC staining in D was completed on a six-week-old pig. IHC staining in F was completed on a nine-week-old pig. Abbreviations: IHC (immunohistochemistry); ILC (innate lymphoid cell); ISH ( in situ hybridization); logFC (log fold-change); No Sig (no significance); PBMC (peripheral blood mononuclear cell); PP (Peyer’s patch); scRNA-seq (single-cell RNA sequencing); t-SNE (t-distributed stochastic neighbor embedding); TCR (T cell receptor)

Article Snippet: A panel to identify γδ, CD4 αβ, and CD8 αβ T cells (gating strategy shown in Supplement to Figure 4-Figure 2B ) included Fixable Viability Dye-eFluor TM 780 (ThermoFisher 65-0865-14); mouse α-pig CD3ε-PE-Cy7 (BD 561477); mouse α-pig γδTCR-iFluor594 (primary antibody Washington State University PG2032; custom conjugation to iFluor594 performed by Caprico Biotechnologies); mouse α-pig CD4-PerCP-Cy5.5 (BD 561474); and mouse α-pig CD8β-PE (BioRad MCA5954PE).

Techniques: Derivative Assay, Flow Cytometry, Immunohistochemistry, Staining, In Situ Hybridization, RNA Sequencing Assay

Journal: bioRxiv

Article Title: Porcine intestinal innate lymphoid cells and lymphocyte spatial context revealed through single-cell RNA sequencing

doi: 10.1101/2022.01.09.475571

Figure Lengend Snippet: (A) Flow cytometry gating strategy used to identify CD2 + Lin - (Lin - = CD172α - CD3ε - CD79α - ) within total viable cells of porcine ileal samples. Gating is shown for a whole ileum sample (containing both regions with and without Peyer’s patches) for pig A (corresponding to pig IDs in C ). (B) Flow cytometry forward- and side-scatter plots of total viable cells (left) and CD2 + Lin - cells (right) within a sample of porcine whole ileum shown in A . A gate identifying cells with scatter profiles consistent with lymphocytes is shown, with percentages of cells within the lymphocyte gate listed in the top right of each plot. (C) Histogram of the percentage of CD45 + cells within CD2 + Lin - cells identified from samples of porcine ileum using the flow cytometry gating strategy shown in A . A fluorescence-minus-one (FMO) sample lacking α-CD45 antibody staining was used as a negative control. (D) Dual fluorescent staining of CD3ε protein and ITGAE RNA in villi (epithelium + lamina propria) of porcine ileum. Left column: overlay of all stains, including CD3ε protein (green), ITGAE RNA (magenta), and nuclei (DAPI staining; blue). Additional columns show individual stain overlays in white, including (from left to right) nuclei, CD3ε protein, and ITGAE RNA. Panels of two separate villi are shown in each row. Panels were selected from larger stitched images as shown in Supplement to Figure 5-Figure 2A . Yellow arrows indicate location of ITGAE + CD3ε - cells. (E) Dual fluorescent staining of CD3ε protein and IL22 RNA in lamina propria/GALT of porcine ileum. Left column: overlay of all stains, including CD3ε protein (green), IL22 RNA (magenta), and nuclei (DAPI staining; blue). Additional columns show individual stain overlays in white, including (from left to right) nuclei, CD3ε protein, and IL22 RNA. Panels of four separate tissue locations are shown in each row. Panels were selected from larger stitched images as shown in Supplement to Figure 5-Figure 2B . Yellow arrows indicate location of IL22 + CD3ε - cells. Orange arrows indicate location of IL22 + CD3ε + cells. Flow cytometry experiments shown in A-C were conducted using three six-week-old pigs. Dual IF/ISH experiments shown in D-E were conducted using a seven-week-old pig used for ileum scRNA-seq. Abbreviations: FMO (fluorescence-minus-one); FSC-A (forward scatter area); FSC-H (forward scatter height); GALT (gut-associated lymphoid tissue); IF (immunofluorescence); ISH ( in situ hybridization); ILC (innate lymphoid cell); scRNA-seq (single-cell RNA sequencing); SSC-A (side scatter area)

Article Snippet: A panel to identify γδ, CD4 αβ, and CD8 αβ T cells (gating strategy shown in Supplement to Figure 4-Figure 2B ) included Fixable Viability Dye-eFluor TM 780 (ThermoFisher 65-0865-14); mouse α-pig CD3ε-PE-Cy7 (BD 561477); mouse α-pig γδTCR-iFluor594 (primary antibody Washington State University PG2032; custom conjugation to iFluor594 performed by Caprico Biotechnologies); mouse α-pig CD4-PerCP-Cy5.5 (BD 561474); and mouse α-pig CD8β-PE (BioRad MCA5954PE).

Techniques: Flow Cytometry, Fluorescence, Staining, Negative Control, Immunofluorescence, In Situ Hybridization, RNA Sequencing Assay

Journal: Molecular pharmaceutics

Article Title: In Situ Vaccination with Cowpea vs Tobacco Mosaic Virus against Melanoma

doi: 10.1021/acs.molpharmaceut.8b00316

Figure Lengend Snippet: Antibodies Used for Flow Cytometry

Article Snippet: Flow cytometry analyses were performed on a BD LSR II flow cytometer (BD Biosciences), and data was analyzed using the FlowJo software (Tree Star Inc.). table ft1 table-wrap mode="anchored" t5 Table 1. caption a7 dye marker vendor innate panel catalog no. clone/host isotype concentration pacific blue CD4S BioLegend 103126 30-F11 Rat IgG2b, K 0.5 mg/mL FITC CD11b BioLegend 10120S Ml/70 Rat IgG2b, K 0.5 mg/mL PE CD80 BioLegend 104707 16–10A1A Hamster IgG 0.2 mg/mL PE-Cy7 CD86 BioLegend 10S014 GL-1 Rat IgG2a, K 0.2 mg/mL APC I-A/I-E (=MHCII) BioLegend 107613 MS/114.1S.2 Rat IgG2b, K 0.2 mg/mL APC/Cy7 Ly-6G BioLegend 127623 1A8 Rat IgG2a, K 0.2 mg/mL APC CD11c BioLegend 10120S N418 A Hamster 0.2 mg/mL PE F4/80 BioLegend 123109 BM8 Rat IgG2a, K 0.2 mg/mL PE/Cy7 Ly6C BioLegend 128018 HK1.4 Rat IgG2c, K 0.2 mg/mL BV60S NK1.1 BioLegend 108737 PK136 IgG2a, k 0.2 mg/mL dye marker vendor adaptive panel catalog no. clone/host isotype concentration pacific blue CD45 BioLegend 103126 30-F11 Rat IgG2b, K 0.5 mg/mL FITC CD4 BioLegend 100405 GK1.5 Rat IgG2b, K 0.5 mg/mL PE CD44 BioLegend 103007 1M7 Rat IgG2b, K 0.2 mg/mL PE-Cy7 CD62L (L-selectin) BioLegend 104417 MEL-14 Rat IgG2a, K 0.2 mg/mL APC CD8 α BioLegend 100712 S3–6.7 Rat IgG2a, K 0.2 mg/mL APC/Cy7 CD3 ε BioLegend 100330 14S-2C11 A. Hamster IgG 0.2 mg/mL Open in a separate window Antibodies Used for Flow Cytometry

Techniques: Marker, Concentration Assay

Journal: Molecular pharmaceutics

Article Title: In Situ Vaccination with Cowpea vs Tobacco Mosaic Virus against Melanoma

doi: 10.1021/acs.molpharmaceut.8b00316

Figure Lengend Snippet: Antibodies Used for Immunohistochemistry

Article Snippet: Flow cytometry analyses were performed on a BD LSR II flow cytometer (BD Biosciences), and data was analyzed using the FlowJo software (Tree Star Inc.). table ft1 table-wrap mode="anchored" t5 Table 1. caption a7 dye marker vendor innate panel catalog no. clone/host isotype concentration pacific blue CD4S BioLegend 103126 30-F11 Rat IgG2b, K 0.5 mg/mL FITC CD11b BioLegend 10120S Ml/70 Rat IgG2b, K 0.5 mg/mL PE CD80 BioLegend 104707 16–10A1A Hamster IgG 0.2 mg/mL PE-Cy7 CD86 BioLegend 10S014 GL-1 Rat IgG2a, K 0.2 mg/mL APC I-A/I-E (=MHCII) BioLegend 107613 MS/114.1S.2 Rat IgG2b, K 0.2 mg/mL APC/Cy7 Ly-6G BioLegend 127623 1A8 Rat IgG2a, K 0.2 mg/mL APC CD11c BioLegend 10120S N418 A Hamster 0.2 mg/mL PE F4/80 BioLegend 123109 BM8 Rat IgG2a, K 0.2 mg/mL PE/Cy7 Ly6C BioLegend 128018 HK1.4 Rat IgG2c, K 0.2 mg/mL BV60S NK1.1 BioLegend 108737 PK136 IgG2a, k 0.2 mg/mL dye marker vendor adaptive panel catalog no. clone/host isotype concentration pacific blue CD45 BioLegend 103126 30-F11 Rat IgG2b, K 0.5 mg/mL FITC CD4 BioLegend 100405 GK1.5 Rat IgG2b, K 0.5 mg/mL PE CD44 BioLegend 103007 1M7 Rat IgG2b, K 0.2 mg/mL PE-Cy7 CD62L (L-selectin) BioLegend 104417 MEL-14 Rat IgG2a, K 0.2 mg/mL APC CD8 α BioLegend 100712 S3–6.7 Rat IgG2a, K 0.2 mg/mL APC/Cy7 CD3 ε BioLegend 100330 14S-2C11 A. Hamster IgG 0.2 mg/mL Open in a separate window Antibodies Used for Flow Cytometry

Techniques:

Journal: Cell reports

Article Title: Molecular, metabolic, and functional CD4 T cell paralysis in the lymph node impedes tumor control

doi: 10.1016/j.celrep.2023.113047

Figure Lengend Snippet: (A) On day 5 after PyMG administration, mice received isotype control, anti-CD3 alone, anti-CD28 alone, or dual anti-CD3 and anti-CD28 antibodies. Cell proliferation of CD4 SMARTA T TS cells in dLN was analyzed on day 8. (B and C) FoxP3-DTR mice were treated on days 0 and 2 with DT, and on days 0, 2, and 5 with CTLA4-blocking or isotype antibodies.

Article Snippet: InVivoMab anti-mouse CD3 ε (Clone 145-2C11) , BioXcell , Cat# BE0001-1, RRID:AB_1107634.

Techniques: Control, Blocking Assay, Expressing, Ex Vivo, MANN-WHITNEY

Journal: Cell reports

Article Title: Molecular, metabolic, and functional CD4 T cell paralysis in the lymph node impedes tumor control

doi: 10.1016/j.celrep.2023.113047

Figure Lengend Snippet:

Article Snippet: InVivoMab anti-mouse CD3 ε (Clone 145-2C11) , BioXcell , Cat# BE0001-1, RRID:AB_1107634.

Techniques: Control, Virus, Recombinant, Staining, Cell Isolation, Blocking Assay, Negative Control, Cloning, Generated, Sequencing, Plasmid Preparation, Transgenic Assay, Software

Journal: Journal of visualized experiments : JoVE

Article Title: A Mouse Model for the Transition of Streptococcus pneumoniae from Colonizer to Pathogen upon Viral Co-Infection Recapitulates Age-Exacerbated Illness

doi: 10.3791/64419

Figure Lengend Snippet: The lungs were harvested, and the immune cell influx was determined by flow cytometry. The representative gating strategy of the different cell types is shown. (A) CD45+, live single cells were gated on and the percentages of (B) PMNs (Ly6G+, CD11b+), macrophages (Ly6G−, Ly6C−, F480+), and monocytes (Ly6G−, Ly6C+), (C) DCs (Ly6G−, CD11c+) and NK cells (NK1.1+, CD3−), (D) TCR− γΔ and CD8 (CD8+, TCRβ+) and CD4 (CD4+, TCRβ+) T cells were determined. Abbreviations: SSC-A = side scatter-peak area; FSC-A = forward scatter-peak area; FSC-H = forward scatter-peak height; SSC-W = side scatter-peak width; L/D = live/dead; FMO = fluorescent minus one; NK = natural killer; PMN = polymorphonuclear leukocyte; DC = dendritic cell; TCR = T cell receptor.

Article Snippet: CD3 ε AF 488 , BD Bioscience , OB153030 , Clone: 145-2C11 DF 1:200.

Techniques: Flow Cytometry

Journal: Journal of visualized experiments : JoVE

Article Title: A Mouse Model for the Transition of Streptococcus pneumoniae from Colonizer to Pathogen upon Viral Co-Infection Recapitulates Age-Exacerbated Illness

doi: 10.3791/64419

Figure Lengend Snippet: Table of Materials

Article Snippet: CD3 ε AF 488 , BD Bioscience , OB153030 , Clone: 145-2C11 DF 1:200.

Techniques: Flow Cytometry, Sterility, Modification, Staining, Blocking Assay, Ointment, Infection

Journal: Journal of visualized experiments : JoVE

Article Title: A Mouse Model for the Transition of Streptococcus pneumoniae from Colonizer to Pathogen upon Viral Co-Infection Recapitulates Age-Exacerbated Illness

doi: 10.3791/64419

Figure Lengend Snippet: Antibody panel 1.

Article Snippet: CD3 ε AF 488 , BD Bioscience , OB153030 , Clone: 145-2C11 DF 1:200.

Techniques: Blocking Assay

Journal: Journal of visualized experiments : JoVE

Article Title: A Mouse Model for the Transition of Streptococcus pneumoniae from Colonizer to Pathogen upon Viral Co-Infection Recapitulates Age-Exacerbated Illness

doi: 10.3791/64419

Figure Lengend Snippet: Antibody panel 2.

Article Snippet: CD3 ε AF 488 , BD Bioscience , OB153030 , Clone: 145-2C11 DF 1:200.

Techniques: Blocking Assay