Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Flow cytometer setup, fluorochrome panel, and labeling.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Flow Cytometry, Labeling, Biomarker Discovery

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Fluorochrome compensation panel graph by sample type and tissue.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Marker

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Gating strategy. Peripheral blood sequential gating strategy for a panel of six antibodies to identify the different cell subpopulations after doublet exclusion. CD45, a pan-leukocyte marker, and CD3, a T-lymphocyte specific marker, were used to define the T-lymphocyte population, with posterior separation of CD4 + and CD8 + cells, CD4 + CD8 + double-positive T cells, and subsequent regulatory CD25 + FoxP3 + and effector CD25 − FoxP3 − cells. Red histograms from unstained control samples and colored histograms from single-stained control samples were used to define the sequential gating, along with gray histograms from fluorescence minus one (FMO) controls to gate for rare cells (CD25 + FoxP3 + ).

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Marker, Control, Staining, Fluorescence

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: CD4/CD8 ratio in the blood (A) , lymph node (B) , and bone marrow (C) of healthy [control group (CG)], sick (M0), and treated dogs (M1, M2, and M3). Results of 22 dogs are represented by mean values ± SEM. The non-parametric Kruskal–Wallis test (one-way ANOVA on ranks) with Dunn's post hoc test was used for statistical comparisons between treatment groups and the CG. The repeated measures ANOVA test with Tukey's post hoc test was used for statistical comparisons inside each treatment group. p -values are indicated in every statistically significant comparison.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Control, Comparison

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Frequency of CD4 + CD8 + double-positive (dp) T cells. The frequency of dp T cells (A–C) expressing CD25 (D–F) and CD25 and FoxP3 (G–I) was evaluated in the peripheral blood (A,D,G) , lymph node (B,E,H) , and bone marrow (C,F,I) of healthy [control group (CG)], sick (M0), and treated dogs (M1, M2, and M3). Results of 22 dogs are represented by box and whisker plots and median, minimum, and maximum values. The non-parametric Kruskal–Wallis test (one-way ANOVA on ranks) with Dunn's post hoc test was used for statistical comparisons between treatment groups and the CG. The repeated measures ANOVA test with Tukey's post hoc test was used for statistical comparisons inside each treatment group. p -values are indicated in every statistically significant comparison.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Expressing, Control, Whisker Assay, Comparison

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Frequency of CD4 + (A) , CD8 + (B) , regulatory (CD25 + FoxP3 + ) (C–F) , and effector (CD4 + CD25 − FoxP3 − /CD8 + CD25 − FoxP3 − ) (G,H) T lymphocytes in the blood of healthy [control group (CG)], sick (M0), and treated dogs (M1, M2, and M3). Results of 22 dogs are represented by box and whisker plots and median, minimum, and maximum values. The non-parametric Kruskal–Wallis test (one-way ANOVA on ranks) with Dunn's post hoc test was used for statistical comparisons between treatment groups and the CG. The repeated measures ANOVA test with Tukey's post hoc test was used for statistical comparisons inside each treatment group. p -values are indicated in every statistically significant comparison.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Control, Whisker Assay, Comparison

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Frequency of CD4 + (A) , CD8 + (B) , regulatory (CD25 + FoxP3 + ) (C–F) , and effector (CD4 + CD25 − FoxP3 − /CD8 + CD25 − FoxP3 − ) (G,H) T lymphocytes in the lymph node of healthy [control group (CG)], sick (M0), and treated dogs (M1, M2, and M3). Results of 22 dogs are represented by box and whisker plots and median, minimum, and maximum values. The non-parametric Kruskal–Wallis test (one-way ANOVA on ranks) with Dunn's post hoc test was used for statistical comparisons between treatment groups and the CG. The repeated measures ANOVA test with Tukey's post hoc test was used for statistical comparisons inside each treatment group. p -values are indicated in every statistically significant comparison.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Control, Whisker Assay, Comparison

Journal: Frontiers in Veterinary Science

Article Title: Immunophenotyping of Peripheral Blood, Lymph Node, and Bone Marrow T Lymphocytes During Canine Leishmaniosis and the Impact of Antileishmanial Chemotherapy

doi: 10.3389/fvets.2020.00375

Figure Lengend Snippet: Frequency of CD4 + (A) , CD8 + (B) , regulatory (CD25 + FoxP3 + ) (C–F) , and effector (CD4 + CD25 − FoxP3 − /CD8 + CD25 − FoxP3 − ) (G,H) T lymphocytes in the bone marrow of healthy [control group (CG)], sick (M0), and treated dogs (M1, M2, and M3). Results of 22 dogs are represented by box and whisker plots and median, minimum, and maximum values. The non-parametric Kruskal–Wallis test (one-way ANOVA on ranks) with Dunn's post hoc test was used for statistical comparisons between treatment groups and the CG. The repeated measures ANOVA test with Tukey's post hoc test was used for statistical comparisons inside each treatment group. p -values are indicated in every statistically significant comparison.

Article Snippet: Cell suspensions (50 μl) were incubated with the following monoclonal antibodies (30 min at 4°C in the dark): rat anti-dog CD45 (clone YKIX716.13, eBioscience Inc.), mouse anti-dog CD3 (clone CA17.2A12, AbD Serotec, UK), anti-dog CD4 (clone YKIX302.9, eBioscience Inc.), rat anti-dog CD8 (clone YCATE55.9, AbD Serotec), and mouse anti-dog CD25 (clone P4A10, eBioscience Inc.) ( ).

Techniques: Control, Whisker Assay, Comparison

Journal: Frontiers in Immunology

Article Title: Effective Activation and Expansion of Canine Lymphocytes Using a Novel Nano-Sized Magnetic Beads Approach

doi: 10.3389/fimmu.2021.604066

Figure Lengend Snippet: The effect of culture temperature on activation of canine CD4 + T lymphocytes. Quantification of activated CD4 + CD25 + T lymphocytes after 24 h (A) and 72 h (B) post stimulation using different MicroBeads concentrations (as indicated) and ConA. To assess the impact of temperature PBLs were incubated in 33°C, 37°C, 38.5°C, 40°C or 41°C in humidified incubator, 5% CO2 (Sanyo Electric). Data are shown as the mean of five dogs (n=5), and error bars indicate SEM. Statistical analysis was performed by One-way analysis of variance (ANOVA) with Tukey’s Multiple Comparison Test (*p < 0.05 ,**p < 0.01, ***p < 0.001).

Article Snippet: To analyze expression level of surface molecules and activation markers the following antibodies were used: anti-canine CD4-APC (clone YKIX302.9), anti-canine CD5-PerCP (clone YKIX322.3), anti-canine CD8α-v450 (clone YCATE55.9) and anti-canine CD25-FITC (clone P4A10) (all purchased from eBioscience™, Thermo Fisher Scientific, Waltham, USA), according to the manufacturer’s recommendation.

Techniques: Activation Assay, Incubation