Journal: Oncology Letters

Article Title: Survival analysis with regard to PD-L1 and CD155 expression in human small cell lung cancer and a comparison with associated receptors

doi: 10.3892/ol.2019.9910

Figure Lengend Snippet: Immunofluorescence double staining of PD-1/TIGIT and CD8 in small cell lung cancer. Nuclear staining with DAPI (blue); CD8 staining with TRITC-goat anti-rabbit second antibody (red) or FITC-donkey anti-rabbit second antibody (green); PD-1 staining with FITC-goat anti-mouse second antibody (green); TIGIT staining with TRITC-donkey anti-goat second antibody (red). Sections were photographed at magnification, ×400. CD, cluster of differentiation; PD, programmed death; TIGIT, T cell immunoreceptor with immunoglobulin and ITIM domains; FITC, fluorescein isothiocyanate; TRITC, tetramethylrhodamine; TILs, tumor-infiltrating lymphocytes.

Article Snippet: Sections were incubated with primary anti-TIGIT antibody and anti-CD8 antibody (1:75 dilution; cat. no. 17335-1-AP; ProteinTech Group, Inc.), or with anti-PD-1 antibody and anti-CD8 antibody (1:75 dilution; cat. no. 17335-1-AP; ProteinTech Group, Inc.) at 4°C.

Techniques: Immunofluorescence, Double Staining, Staining

Journal: Nature Communications

Article Title: Th1-poised naive CD4 T cell subpopulation reflects anti-tumor immunity and autoimmune disease

doi: 10.1038/s41467-025-57237-3

Figure Lengend Snippet: a Representative histogram of IL-7R, Adgre5, CD5 expression in IL-18R/Ly6C double-positive (DP) and double-negative (DN) naive CD4 T cells. b Representative dot plot of IL-18R and Ly6C expression in IL-7R sup-hi and IL-7R low naive CD4 T cells and memory-phenotype CD4 T cells (MP). c Representative dot plot of IL-18R/Ly6C DP population in naive CD4 T cells from each indicated tissue. d Tissue distribution of IL-18R/Ly6C DP population in naive CD4 T cells ( n = 10). e Representative histogram of naive CD4 T cells from spleen and thymus. f UMAP of murine naïve CD4 T cells from two different tissues (thymic CD4 SP and spleen). Cells were colored by cluster identity using label transfer of naïve CD4 T cell clusters from Fig. . g Stacked bar graph showing the proportion of naïve CD4 T cell clusters from thymus (Thy) and spleen (Spl). h Dot plots showing the distribution of pseudotime for the single cells in each cluster. i Alluvial plot to track the clonal population in thymus (Thy) and spleen (Spl) using TCR-seq data. j Box plots for the Shannon diversity index of TCR repertoire for each cluster in each tissue. k Expression of Ly6C and IL-18R in naive CD4 and CD8 single-positive (SP) cells in thymocytes stimulated with anti-CD3 for 3 days. l Proportion of Ly6C + cells in SP CD4 naive thymocytes stimulated by the indicated cytokines for 3 days ( n = 5). Statistical significance was confirmed by two-sided Mann-Whitney U -test with control group and each experimental group ( p = 0.0079). m Proportion of Ly6C + naive CD4 T cells in the splenocyte of WT, Stat1 -/- , Ifnar -/- , and Ifnar -/- Ifngr -/- mice (WT n = 5, Stat1 KO n = 5, Ifnar1 KO n = 3, Ifnar1 Ifngr1 KO n = 2). Data presented as the mean ± S.D. Statistical significance was confirmed by the two-sided Mann-Whitney U -test (* p < 0.05, ** p < 0.01, *** p < 0.001). Source data and exact statistics value are provided as a Source Data file.

Article Snippet: CD8 and NK depletion was performed by a single i.p injection of 200μg of anti-CD8 antibody (clone 2.43, BioXcell, #BE0061, lot 811521N1) and anti-NK1.1 antibody (clone PK136, BioXcell, #BE0036 lot 828622A2).

Techniques: Expressing, MANN-WHITNEY, Control

Journal: Nature Communications

Article Title: Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models

doi: 10.1038/s41467-021-26830-7

Figure Lengend Snippet: a – e 1-week-old normal dogs were either co-injected with AAV.CK8.SpCas9 and AAV.RSV.AP or injected with AAV.RSV.AP only. a Representative HE, AP, CD4, and CD8 stainings. b Cas9 , and AP vector genome quantification (N, n = 5; others, n = 3). c Serum Cas9 antibody (Naive, n = 8; 6wks., n = 1). d Cas9 ELISpot assay on lymphocytes (Naive, n = 5; 6wks., n = 3). e Muscle cytokine transcript quantification (Naive, n = 8; others, n = 3). f - j Similar to a - e except 1-m-old dogs were injected. f Representative HE, AP, CD4, and CD8 stainings. Arrowhead, T-cell infiltration wiped out AP expression. g Cas9 and AP vector genome quantification ( N , n = 4; others, n = 3). h Serum Cas9 antibody ( N , n = 12; 6wks., n = 1). i Cas9 ELISpot assay on lymphocytes (Naive, n = 5; 6wks., n = 1). j Muscle cytokine transcript quantification ( N , n = 14; others, n = 3). k – p Similar to a - e except adult dogs were injected with either AAV.CK8.SpCas9 or AAV.RSV.AP. k Representative HE, AP, CD8, and granzyme B stainings. The boxed region was magnified. Yellow arrowhead, a dying myofiber lost dystrophin expression and infiltrated with CD8 + and granzyme B + T cells. l CD8 + T-cell quantification ( n = 3 for all categories). m Dystrophin and Cas9 western blot. n Serum Cas9 antibody (Naive, n = 27; others, n = 3). o Cas9 ELISpot assay on lymphocytes (Naive, n = 6; others, n = 3). p Muscle cytokine transcript quantification from adult dogs ( N , n = 12; others, n = 3). Immune cells are stained in dark brown, and AP is stained in blue. AP, alkaline phosphatase. N, No AAV. N/A, non-applicable. wks., weeks post-injection. Data are mean ± SEM. Statistical analysis was performed using Crawford-Howell test for c , h , i , One-way ANOVA with Tukey’s multiple comparisons for b , e , g , j , l , and n - p and Student’s t-test for d . See source data file for the exact p -value. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Rat monoclonal IgG1 antibody against canine CD8 (Catalog# MCA1039, clone YCATE 55.9, Lot # 149349, 1:200, Bio-Rad Laboratories, Inc.) was used to detect CD8 + T and was visualized using a biotin-conjugated goat anti-rat IgG (H + L) antibody (A10517, Lot # 1441195, 1:1000, ThermoFisher Scientific).

Techniques: Injection, Plasmid Preparation, Enzyme-linked Immunospot, Expressing, Western Blot, Staining

Journal: Nature Communications

Article Title: Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models

doi: 10.1038/s41467-021-26830-7

Figure Lengend Snippet: a Representative HE, dystrophin, CD4, and CD8 stainings from biopsy. b , Representative dystrophin and CD8 stainings from necropsied muscles. c Representative HE, dystrophin, CD8, and granzyme B staining from a necropsied muscle. d Cas9 and gRNA vector genome quantification (N, n = 7; 3wks., n = 1; 6wks., n = 11). e Cas9 transcript quantification ( N , n = 7; 3wks., n = 1; 6wks., n = 11). f Serum Cas9 antibody ( N , n = 16, 8, 9, 11, 12, 23, 11 and 18 for 0, 1, 2, 3, 4, 6, 8, and 10 weeks after birth, respectively; dog#1, n = 1 for 0, 4–10 weeks after birth). g PBMC IFN-γ ELISPOT against Cas9 ( N , n = 11; dog #1 n = 1 for all-time points). h Representative HE, dystrophin, CD4, and CD8 staining from biopsy. i Representative dystrophin and CD8 staining from necropsied muscles. j Representative HE, dystrophin, CD8, and granzyme B stainings from a necropsied muscle. k Cas9 and gRNA vector genome quantification ( N , n = 7; 3wks., n = 1; 6wks., n = 18). l Cas9 transcript quantification ( N , n = 7, 3wks., n = 1; 6wks., n = 18). m Serum Cas9 antibody ( N , n = 16, 8, 9, 11, 12, 23, 11 and 18 for 0, 1, 2, 3, 4, 6, 8, and 16 weeks after birth, respectively; dog#2, n = 1 for 0, 4–16 weeks after birth). n Cas9 specific IFN-γ ELISpot assay on PBMCs ( N , n = 11; dog #1 n = 1 for all-time points). o Representative HE, dystrophin, CD4 and CD8 staining from the control dog that received the micro-dystrophin vector. p Muscle cytokine transcript quantification from all three dogs ( N , n = 7;dog#1, 3wks. n = 1, 6wks. n = 11; dog#2, 3wks. n = 1, 12wks. n = 18; μDys injected dog, 6wks. n = 1, 24wks. n = 1). Immune cells are stained in dark brown. Arrow, a granzyme B-positive T cell. N , No AAV. wks., weeks post-injection. Data are mean ± SEM. Statistical analysis was performed using Crawford–Howell test for d – g and k – n , One-way ANOVA with Tukey’s multiple comparisons for p . See source data file for the exact p -value. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Rat monoclonal IgG1 antibody against canine CD8 (Catalog# MCA1039, clone YCATE 55.9, Lot # 149349, 1:200, Bio-Rad Laboratories, Inc.) was used to detect CD8 + T and was visualized using a biotin-conjugated goat anti-rat IgG (H + L) antibody (A10517, Lot # 1441195, 1:1000, ThermoFisher Scientific).

Techniques: Muscles, Staining, Plasmid Preparation, Enzyme-linked Immunospot, Control, Injection

Journal: Nature Communications

Article Title: Cas9-specific immune responses compromise local and systemic AAV CRISPR therapy in multiple dystrophic canine models

doi: 10.1038/s41467-021-26830-7

Figure Lengend Snippet: a Representative HE, AP CD4, and CD8 stainings. b Cas9 and AP vector genome quantification ( N , n = 7; others, n = 1). c Cas9 and AP transcript quantification ( N , n = 7; others, n = 1). d AP + myofiber quantification ( n = 1 in all categories). e Serum Cas9 antibody ( N , n = 16, 8, 9, 11, 12, 23, 11 and 18 for 0, 1, 2, 3, 4, 6, 8, and 16 weeks after birth, respectively; dog#1, n = 1 for all-time points). f Cas9-specific IFN-γ ELISpot assay on PBMCs ( N , n = 11; dog #1 n = 1 for all-time points). g Representative HE, AP, CD4, and CD8 staining. h Cas9 and AP vector genome quantification ( N , n = 7; others, n = 1). i Cas9 and AP transcript quantification (in both panels: N , n = 7; each other group, n = 1). j AP + myofiber quantification ( n = 1 in each group). k Serum Cas9 antibody ( N , n = 16, 8, 9, 11, 12, 23, 11 and 18 for 0, 1, 2, 3, 4, 6, 8, and 16 weeks after birth, respectively; dog#2, n = 1 for all time points). l , PBMC IFN-γ ELISpot against Cas9 ( N , n = 11; dog #1 n = 1 for all time points). m Representative HE, AP, CD4, and CD8 stainings. n AP vector genome quantification ( N , n = 7; others, n = 1). o AP transcript quantification ( N , n = 7; others, n = 1). p Muscle cytokine transcript quantification from all 3 dogs at the indicated time points post-injection ( N , n = 14; dog#1, 3wks. n = 1, 6wks. n = 1, 12wks. n = 1; dog#2, 3wks. n = 1, 6wks. n = 1, 12wks. n = 1; dog injected with AP vector only, 6wks. n = 1, 52 wks. n = 1). AP, alkaline phosphatase. N, No AAV. wks., weeks post-injection. Data are mean ± SEM. Statistical analysis was performed using Crawford-Howell test for b , c , e , f , h , i , k , l , n , o , and One-way ANOVA with Tukey’s multiple comparisons for p . See source data file for exact p -value. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: Rat monoclonal IgG1 antibody against canine CD8 (Catalog# MCA1039, clone YCATE 55.9, Lot # 149349, 1:200, Bio-Rad Laboratories, Inc.) was used to detect CD8 + T and was visualized using a biotin-conjugated goat anti-rat IgG (H + L) antibody (A10517, Lot # 1441195, 1:1000, ThermoFisher Scientific).

Techniques: Plasmid Preparation, Enzyme-linked Immunospot, Staining, Injection

Journal: The Laryngoscope

Article Title: T-Helper 2 Lymphocyte Immunophenotype Is Associated With Iatrogenic Laryngotracheal Stenosis

doi: 10.1002/lary.27321

Figure Lengend Snippet: Lymphocyte immunofluorescence of human normal and iLTS specimens. Photomicrographs depicting little (A) T-lymphocyte (CD3) or (C) helper T-lymphocyte (CD4) presence in normal tracheal mucosa compared with intense staining for (B) CD3 (green chromagen) and (D) CD4 (red chromagen), indicative of a dense CD4 + T-lymphocyte infiltrate within iLTS periepithelial mucosa. There was no evidence of (E, F) cytotoxic T-lymphocytes (CD8) or (G, H) B-lymphocytes (CD20) in normal or diseased specimens. All specimens 400 × magnification. [Color figure can be viewed in the online issue, which is available at www.laryngoscope.com.]

Article Snippet: Immunofluorescence slides underwent primary antibody staining with rabbit anti-human CD 3 (lot #: ab5690, Abcam, Cambridge, MA), mouse anti-human CD4 (lot #: ab846, Abcam), rabbit anti-human CD8 (lot #: sc-7188, Santa Cruz Biotechnology, Dallas, TX), and rabbit anti-human CD20 (lot #: {"type":"entrez-protein","attrs":{"text":"RPA11836","term_id":"1517911547","term_text":"RPA11836"}} RPA11836 , Reprokine Ltd, Rehovot, Israel) followed by secondary antibody (Life Technologies, ThermoFisher Scientific, Waltham, MA).

Techniques: Immunofluorescence, Staining

Journal: The Laryngoscope

Article Title: T-Helper 2 Lymphocyte Immunophenotype Is Associated With Iatrogenic Laryngotracheal Stenosis

doi: 10.1002/lary.27321

Figure Lengend Snippet: Human T-lymphocytes and T-helper lymphocytes are present in significantly greater numbers in iLTS specimens than normal subglottic specimens. Bar graphs comparing the cell density (cells/hpf ) for mean total cells (A), CD3 + cells (B), CD8 + cells (C), CD3 + (D), CD3+/CD4 + cells (E), and CD3+/CD8 + cells (F) in areas of high-density inflammation in both iLTS and normal specimens.* P < 0.05; **P < 0.01. [Color figure can be viewed in the online issue, which is available at www.laryngoscope.com.]

Article Snippet: Immunofluorescence slides underwent primary antibody staining with rabbit anti-human CD 3 (lot #: ab5690, Abcam, Cambridge, MA), mouse anti-human CD4 (lot #: ab846, Abcam), rabbit anti-human CD8 (lot #: sc-7188, Santa Cruz Biotechnology, Dallas, TX), and rabbit anti-human CD20 (lot #: {"type":"entrez-protein","attrs":{"text":"RPA11836","term_id":"1517911547","term_text":"RPA11836"}} RPA11836 , Reprokine Ltd, Rehovot, Israel) followed by secondary antibody (Life Technologies, ThermoFisher Scientific, Waltham, MA).

Techniques:

Journal: The Laryngoscope

Article Title: T-Helper 2 Lymphocyte Immunophenotype Is Associated With Iatrogenic Laryngotracheal Stenosis

doi: 10.1002/lary.27321

Figure Lengend Snippet: Flow cytometry of mouse tracheas demonstrates a CD4 + T-lymphocyte predominance. Cell surface protein expression with (A) increased CD3 + expression at days 4, 7, and 10 in the mouse iLTS group when compared to controls. Subset analysis revealed the T-lymphocytes to be (B) CD3+/CD4 + T-helper cells with few (C) CD3+/CD8 + cytotoxic T-lymphocytes. [Color figure can be viewed in the online issue, which is available at www.laryngoscope.com.]

Article Snippet: Immunofluorescence slides underwent primary antibody staining with rabbit anti-human CD 3 (lot #: ab5690, Abcam, Cambridge, MA), mouse anti-human CD4 (lot #: ab846, Abcam), rabbit anti-human CD8 (lot #: sc-7188, Santa Cruz Biotechnology, Dallas, TX), and rabbit anti-human CD20 (lot #: {"type":"entrez-protein","attrs":{"text":"RPA11836","term_id":"1517911547","term_text":"RPA11836"}} RPA11836 , Reprokine Ltd, Rehovot, Israel) followed by secondary antibody (Life Technologies, ThermoFisher Scientific, Waltham, MA).

Techniques: Flow Cytometry, Expressing