Journal: Frontiers in Immunology

Article Title: A Unique Cellular and Molecular Microenvironment Is Present in Tertiary Lymphoid Organs of Patients with Spontaneous Prostate Cancer Regression

doi: 10.3389/fimmu.2017.00563

Figure Lengend Snippet: IL17 production, B cell activation, and CD8 T cell accumulation are evident in prostate tumor-associated tertiary lymphoid organs (TLO) . The 5-μm thick paraffin sections were stained with antibodies against proliferating cell nuclear antigen (PCNA, red), CD8 (green), and CD20 (white) to identify B cells and CD8 T cells in tumor-associated TLO. Serial sections were stained with antibodies against CD21 (red), IL17 (green), and CD3 (white). Representative 200× magnification pictures from the same TLO areas showed in H&E stains were taken with a Zeiss Axioplan Microscope and recorded with a Hamamatsu Camera. (A,E) Well-populated B cells follicles with small numbers of small proliferating B cells, small follicular dendritic cell (FDC) networks, and intrafollicular CD8 T cells are appreciated in prostatic intraepithelial neoplasia (PIN) prostatectomy specimens. (B,F) Large PCNA + CD20 + B blasts, and CD8 T cells interspersed inside B cell follicles are detected in prostate samples from patients at intermediate stages of cancer. (C,G) A germinal center with a concentric FDC network, containing multiple large proliferating B blasts and surrounded by CD8 T cells was found inside some TLO of prostatectomy specimens from patients afflicted by advanced cancer. (D,H) Although proliferating B cells are notably reduced, B cell and CD8 T cells are still organized in TLO from evanescent prostate cancer patients. Yellow arrows point to proliferating B cells, while white arrow is depicting a proliferating CD8 T cell. Scale bars represent 100 μm.

Article Snippet: The primary antibodies were as follows: goat anti-human CD105 (AF1097, R&D Systems), rabbit anti cyclooxygenase 2 (GTX15191, GeneTex), mouse anti-human CD68 (clone PG-M1, GeneTex), goat anti-CD3 epsilon (clone M-20, Santa Cruz Biotechnology), rabbit anti-T bet (H-210, Santa Cruz biotechnology), rat anti-human Foxp3 (PCH101, eBioscience), goat-anti proliferating cell nuclear antigen (PCNA) (clone C-20, Santa Cruz Biotechnology), mouse anti-human Ki-67 (clone MIB-1, Dakocytomation), rabbit anti-human CD8 (clone SP16, Thermo Fisher Scientific), mouse anti-human CD20 (clone L-26, Abcam), rabbit anti-human PD-L1 (Invitrogen, PA5-28115), rabbit anti-IL17 (H-132, Santa Cruz Biotechnology), mouse anti-human CD21 (clone 2G9, Thermo Fisher Scientific), rat anti-peripheral node addressin (clone MECA-79, BD Pharmigen), rabbit anti-human CD138 (RB-9422-P1, Thermo Fisher Scientific), mouse anti-podoplanin (clone D2-40, GTX31231, GeneTex), rabbit anti-granzyme B (clone EPR8260, Abcam), rat anti-human DC-LAMP (clone 1010E1.01, Novus Biologicals), rabbit anti-human prostate stem cell antigen (PSCA) (GTX15168, GeneTex), rabbit anti-human CXCL10 (GTX31176, GeneTex), biotin-mouse anti-smooth muscle actin (clone 1A4, Thermo Fisher Scientific), and mouse anti-human plasma cell (clone LIV3G11, Thermo Fisher Scientific).

Techniques: Activation Assay, Staining, Microscopy

Journal: Frontiers in Immunology

Article Title: A Unique Cellular and Molecular Microenvironment Is Present in Tertiary Lymphoid Organs of Patients with Spontaneous Prostate Cancer Regression

doi: 10.3389/fimmu.2017.00563

Figure Lengend Snippet: Enumeration of CD8 T cells in tertiary lymphoid organs (TLO) and tumor areas of prostate cancer . CD8 T cells were counted in all TLO contained in individual prostatectomy specimens or in five to eight randomly selected tumor areas lacking TLO (200× magnification). (A) Average number of CD8 T cells inside TLO and (B) average number of CD8 T cells in tumor regions are shown. n = 10–19 measurements/patient cohort. Bars represent mean ± SEM. Statistically significant differences (*** p ≤ 0.0005, **** p < 0.0001) were calculated by using two-tailed Student’s t -test with GraphPad Prism.

Article Snippet: The primary antibodies were as follows: goat anti-human CD105 (AF1097, R&D Systems), rabbit anti cyclooxygenase 2 (GTX15191, GeneTex), mouse anti-human CD68 (clone PG-M1, GeneTex), goat anti-CD3 epsilon (clone M-20, Santa Cruz Biotechnology), rabbit anti-T bet (H-210, Santa Cruz biotechnology), rat anti-human Foxp3 (PCH101, eBioscience), goat-anti proliferating cell nuclear antigen (PCNA) (clone C-20, Santa Cruz Biotechnology), mouse anti-human Ki-67 (clone MIB-1, Dakocytomation), rabbit anti-human CD8 (clone SP16, Thermo Fisher Scientific), mouse anti-human CD20 (clone L-26, Abcam), rabbit anti-human PD-L1 (Invitrogen, PA5-28115), rabbit anti-IL17 (H-132, Santa Cruz Biotechnology), mouse anti-human CD21 (clone 2G9, Thermo Fisher Scientific), rat anti-peripheral node addressin (clone MECA-79, BD Pharmigen), rabbit anti-human CD138 (RB-9422-P1, Thermo Fisher Scientific), mouse anti-podoplanin (clone D2-40, GTX31231, GeneTex), rabbit anti-granzyme B (clone EPR8260, Abcam), rat anti-human DC-LAMP (clone 1010E1.01, Novus Biologicals), rabbit anti-human prostate stem cell antigen (PSCA) (GTX15168, GeneTex), rabbit anti-human CXCL10 (GTX31176, GeneTex), biotin-mouse anti-smooth muscle actin (clone 1A4, Thermo Fisher Scientific), and mouse anti-human plasma cell (clone LIV3G11, Thermo Fisher Scientific).

Techniques: Two Tailed Test

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Costimulatory pathways in multiple sclerosis: distinctive expression of PD-1 and PD-L1 in patients with different patterns of disease.

doi: 10.4049/jimmunol.0901038

Figure Lengend Snippet: FIGURE 1. MBP-stimulated CD4 and CD8 expressing PD-1. Representative results of MBP-stimulated PBMC of patients with either AMS or SMS are shown. Top panels, PD-1-expressing MBP-stimulated CD4 T lymphocytes. Bottom panels, PD-1-expressing MBP-stimulated CD8 T lymphocytes. In the upper right corners, the percentage of CD4/PD-1 and CD8/PD-1 T cells relative to the total number of lymphocytes are indicated. Summary results obtained in AMS and SMS patients are presented in A and B. The boxes stretch from the 25th to the 75th percentile; the lines across the boxes indicate the median values; the lines stretching from the boxes indicate extreme values. Statistical significance is shown.

Article Snippet: The following mAbs were used in this study: anti-human CD4 (clone 13B8.2; mouse IgG2a isotype), anti-human CD14 (clone 116; mouse IgG1 isotype), anti-human CD19 (clone J4.119; mouse IgG1 isotype) coupled to R-PE-Cyanine 5 Tandem (PE-Cy5; Caltag Laboratories); anti-human CD8 (clone SFCI21thy2D3; mouse IgG1 isotype) PE-Cyanin-7 (BeckmanCoulter); anti-human CD86 (clone BU63; mouse IgG1 isotype), antihuman CD80 (clone 3H5; mouse IgG1 isotype; Serotec) coupled to FITC; anti-human PD-L1 (gift from Dr. L. Chen) and anti-mouse IgG (H L chain) coupled to FITC (eBioscience); anti-human-B7H3 (gift from Dr. L. Chen) coupled FITC anti-human PD-1 (clone MIH4) coupled to PE (mouse-IgG1 isotype; eBioscience); and anti-human annexin V coupled to FITC (mouse-IgG1 isotype; Beckman Coulter).

Techniques: Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Costimulatory pathways in multiple sclerosis: distinctive expression of PD-1 and PD-L1 in patients with different patterns of disease.

doi: 10.4049/jimmunol.0901038

Figure Lengend Snippet: FIGURE 4. Annexin V- and PD-1-expressing MBP-stimulated CD4 and CD8 T lymphocytes. Representative results of MBP-stimulated PBMC of patients with either AMS or SMS are shown. Top panels, MBP-stimulated CD4 T lymphocytes expressing annexin V and PD-1. Bottom panels, MBP-stimulated CD8 T lymphocytes expressing annexin V and PD-1. Summary results obtained in AMS and SMS patients are presented in A and B. The boxes stretch from the 25th to the 75th percentile; the lines across the boxes indicate the median values; the lines stretching from the boxes indicate extreme values. Statistical significance is shown.

Article Snippet: The following mAbs were used in this study: anti-human CD4 (clone 13B8.2; mouse IgG2a isotype), anti-human CD14 (clone 116; mouse IgG1 isotype), anti-human CD19 (clone J4.119; mouse IgG1 isotype) coupled to R-PE-Cyanine 5 Tandem (PE-Cy5; Caltag Laboratories); anti-human CD8 (clone SFCI21thy2D3; mouse IgG1 isotype) PE-Cyanin-7 (BeckmanCoulter); anti-human CD86 (clone BU63; mouse IgG1 isotype), antihuman CD80 (clone 3H5; mouse IgG1 isotype; Serotec) coupled to FITC; anti-human PD-L1 (gift from Dr. L. Chen) and anti-mouse IgG (H L chain) coupled to FITC (eBioscience); anti-human-B7H3 (gift from Dr. L. Chen) coupled FITC anti-human PD-1 (clone MIH4) coupled to PE (mouse-IgG1 isotype; eBioscience); and anti-human annexin V coupled to FITC (mouse-IgG1 isotype; Beckman Coulter).

Techniques: Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Costimulatory pathways in multiple sclerosis: distinctive expression of PD-1 and PD-L1 in patients with different patterns of disease.

doi: 10.4049/jimmunol.0901038

Figure Lengend Snippet: FIGURE 5. pAkt-expressing MBP-stimulated CD4 and CD8 T lymphocytes. Representative results of MBP-stimulated PBMC of patients with either AMS or stable SMS are shown. Top panels, MBP-stimulated CD4 T lymphocytes expressing pAkt. Bottom panels, MBP-stimulated CD8 T lymphocytes expressing pAkt. Summary results obtained in AMS and SMS patients are presented in A and B. The boxes stretch from the 25th to the 75th percentile; the lines across the boxes indicate the median values; the lines stretching from the boxes indicate extreme values. Statistical significance is shown.

Article Snippet: The following mAbs were used in this study: anti-human CD4 (clone 13B8.2; mouse IgG2a isotype), anti-human CD14 (clone 116; mouse IgG1 isotype), anti-human CD19 (clone J4.119; mouse IgG1 isotype) coupled to R-PE-Cyanine 5 Tandem (PE-Cy5; Caltag Laboratories); anti-human CD8 (clone SFCI21thy2D3; mouse IgG1 isotype) PE-Cyanin-7 (BeckmanCoulter); anti-human CD86 (clone BU63; mouse IgG1 isotype), antihuman CD80 (clone 3H5; mouse IgG1 isotype; Serotec) coupled to FITC; anti-human PD-L1 (gift from Dr. L. Chen) and anti-mouse IgG (H L chain) coupled to FITC (eBioscience); anti-human-B7H3 (gift from Dr. L. Chen) coupled FITC anti-human PD-1 (clone MIH4) coupled to PE (mouse-IgG1 isotype; eBioscience); and anti-human annexin V coupled to FITC (mouse-IgG1 isotype; Beckman Coulter).

Techniques: Expressing

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Costimulatory pathways in multiple sclerosis: distinctive expression of PD-1 and PD-L1 in patients with different patterns of disease.

doi: 10.4049/jimmunol.0901038

Figure Lengend Snippet: FIGURE 6. Blockade of the PD-1-PD-L1 pathway. pAKT-expressing, MBP-stimulated CD4 and CD8 T lymphocytes.

Article Snippet: The following mAbs were used in this study: anti-human CD4 (clone 13B8.2; mouse IgG2a isotype), anti-human CD14 (clone 116; mouse IgG1 isotype), anti-human CD19 (clone J4.119; mouse IgG1 isotype) coupled to R-PE-Cyanine 5 Tandem (PE-Cy5; Caltag Laboratories); anti-human CD8 (clone SFCI21thy2D3; mouse IgG1 isotype) PE-Cyanin-7 (BeckmanCoulter); anti-human CD86 (clone BU63; mouse IgG1 isotype), antihuman CD80 (clone 3H5; mouse IgG1 isotype; Serotec) coupled to FITC; anti-human PD-L1 (gift from Dr. L. Chen) and anti-mouse IgG (H L chain) coupled to FITC (eBioscience); anti-human-B7H3 (gift from Dr. L. Chen) coupled FITC anti-human PD-1 (clone MIH4) coupled to PE (mouse-IgG1 isotype; eBioscience); and anti-human annexin V coupled to FITC (mouse-IgG1 isotype; Beckman Coulter).

Techniques: Expressing

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: Asian Journal of Pharmaceutical Sciences

Article Title: Lignin-assisted construction of sub-10 nm supramolecular self-assembly for photothermal immunotherapy and potentiating anti-PD-1 therapy against primary and distant breast tumors

doi: 10.1016/j.ajps.2022.07.002

Figure Lengend Snippet: In vivo immune responses assessment. (A) Flow cytometry assay of CD80 + or CD86 + gated on CD11c + in the TDLNs of mice. (B) Flow cytometry assay illustrated the percent of CD8a + T cells and CD4 + T cells gated on CD3 + T cells in splenocytes of mice. (C-D) Quantitative percentages of CD80 + or CD86 + expressions gated on CD11c + according to (A). (E) Quantitative percentages of CD8a + T cells and CD4 + T cells in splenocytes according to (B). (F-H) Quantitative analysis of the immune-associated cytokines in serum, as determined by ELISA (IL-2, TNF-α and IFN-γ). ( n = 3, mean ± SD, * P < 0.05, ** P < 0.01 and ## P < 0.0001).

Article Snippet: Fluorochrome-labeled anti-mouse monoclonal antibodies (CD11c-allophycocyanin (APC), CD80-fluorescein isothiocyanate (FITC), CD86-phycoerythrin (PE), CD3-FITC, CD8a-PE, and CD4-APC) were bought from Proteintech.

Techniques: In Vivo, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Medicine

Article Title: Expression and clinical significance of PD-L1 and infiltrated immune cells in the gastric adenocarcinoma microenvironment

doi: 10.1097/MD.0000000000036323

Figure Lengend Snippet: Correlation between PD-L1 and different immune cells.

Article Snippet: After blocking with 5% bovine serum albumin, the sections were incubated with an rabbit antihuman PD-L1 monoclonal (1:200; clone number E1L3N; CST), mouse antihuman CD3 monoclonal (ready to use; clone number LN10; ZSGB-BIO; China), mouse antihuman CD4 monoclonal (ready to use; clone number UMAB64; ZSGB-BIO; China), rabbit antihuman CD8 monoclonal (ready to use; clone number SP16; ZSGB-BIO; China), mouse antihuman CD11c monoclonal (1:50; clone number 5D11; ZSGB-BIO; China), mouse antihuman CD19 monoclonal (ready to use; clone number UMAB103; ZSGB-BIO; China), mouse antihuman CD31 monoclonal (ready to use; clone number UMAB30; ZSGB-BIO; China), mouse antihuman CD56 monoclonal (ready to use; clone number UMAB83; ZSGB-BIO; China), mouse antihuman CD68 monoclonal (ready to use; clone number KP1 ZSGB-BIO; China), mouse antihuman α-SMA monoclonal (ready to use; clone number UMAB237; ZSGB-BIO; China) at 4 °C overnight.

Techniques:

Journal: Medicine

Article Title: Expression and clinical significance of PD-L1 and infiltrated immune cells in the gastric adenocarcinoma microenvironment

doi: 10.1097/MD.0000000000036323

Figure Lengend Snippet: Survival data were analyzed by PD-L1 expression with different immune cells infiltration. (A, C, E, G, I, K) TPD-L1 with CD3, CD4, CD8, CD19, CD31, and CD68 infiltration. (B, D, F, H, J, L) IPD-L1 with CD3, CD4, CD8, CD19, CD31, and CD68 infiltration.

Article Snippet: After blocking with 5% bovine serum albumin, the sections were incubated with an rabbit antihuman PD-L1 monoclonal (1:200; clone number E1L3N; CST), mouse antihuman CD3 monoclonal (ready to use; clone number LN10; ZSGB-BIO; China), mouse antihuman CD4 monoclonal (ready to use; clone number UMAB64; ZSGB-BIO; China), rabbit antihuman CD8 monoclonal (ready to use; clone number SP16; ZSGB-BIO; China), mouse antihuman CD11c monoclonal (1:50; clone number 5D11; ZSGB-BIO; China), mouse antihuman CD19 monoclonal (ready to use; clone number UMAB103; ZSGB-BIO; China), mouse antihuman CD31 monoclonal (ready to use; clone number UMAB30; ZSGB-BIO; China), mouse antihuman CD56 monoclonal (ready to use; clone number UMAB83; ZSGB-BIO; China), mouse antihuman CD68 monoclonal (ready to use; clone number KP1 ZSGB-BIO; China), mouse antihuman α-SMA monoclonal (ready to use; clone number UMAB237; ZSGB-BIO; China) at 4 °C overnight.

Techniques: Expressing

Journal: Medicine

Article Title: Expression and clinical significance of PD-L1 and infiltrated immune cells in the gastric adenocarcinoma microenvironment

doi: 10.1097/MD.0000000000036323

Figure Lengend Snippet: Univariate analysis of overall survival.

Article Snippet: After blocking with 5% bovine serum albumin, the sections were incubated with an rabbit antihuman PD-L1 monoclonal (1:200; clone number E1L3N; CST), mouse antihuman CD3 monoclonal (ready to use; clone number LN10; ZSGB-BIO; China), mouse antihuman CD4 monoclonal (ready to use; clone number UMAB64; ZSGB-BIO; China), rabbit antihuman CD8 monoclonal (ready to use; clone number SP16; ZSGB-BIO; China), mouse antihuman CD11c monoclonal (1:50; clone number 5D11; ZSGB-BIO; China), mouse antihuman CD19 monoclonal (ready to use; clone number UMAB103; ZSGB-BIO; China), mouse antihuman CD31 monoclonal (ready to use; clone number UMAB30; ZSGB-BIO; China), mouse antihuman CD56 monoclonal (ready to use; clone number UMAB83; ZSGB-BIO; China), mouse antihuman CD68 monoclonal (ready to use; clone number KP1 ZSGB-BIO; China), mouse antihuman α-SMA monoclonal (ready to use; clone number UMAB237; ZSGB-BIO; China) at 4 °C overnight.

Techniques: Expressing

Journal: Medicine

Article Title: Expression and clinical significance of PD-L1 and infiltrated immune cells in the gastric adenocarcinoma microenvironment

doi: 10.1097/MD.0000000000036323

Figure Lengend Snippet: Multivariate analysis of overall survival.

Article Snippet: After blocking with 5% bovine serum albumin, the sections were incubated with an rabbit antihuman PD-L1 monoclonal (1:200; clone number E1L3N; CST), mouse antihuman CD3 monoclonal (ready to use; clone number LN10; ZSGB-BIO; China), mouse antihuman CD4 monoclonal (ready to use; clone number UMAB64; ZSGB-BIO; China), rabbit antihuman CD8 monoclonal (ready to use; clone number SP16; ZSGB-BIO; China), mouse antihuman CD11c monoclonal (1:50; clone number 5D11; ZSGB-BIO; China), mouse antihuman CD19 monoclonal (ready to use; clone number UMAB103; ZSGB-BIO; China), mouse antihuman CD31 monoclonal (ready to use; clone number UMAB30; ZSGB-BIO; China), mouse antihuman CD56 monoclonal (ready to use; clone number UMAB83; ZSGB-BIO; China), mouse antihuman CD68 monoclonal (ready to use; clone number KP1 ZSGB-BIO; China), mouse antihuman α-SMA monoclonal (ready to use; clone number UMAB237; ZSGB-BIO; China) at 4 °C overnight.

Techniques: Expressing

Journal: Acta Neuropathologica

Article Title: The structural differences between patient-derived α-synuclein strains dictate characteristics of Parkinson’s disease, multiple system atrophy and dementia with Lewy bodies

doi: 10.1007/s00401-020-02157-3

Figure Lengend Snippet: Assessment of immune response after inoculation of different patient-derived homogenates and PMCA-amplified αSYN strains in association with rAAV2/7-mediated αSYN expression. a Representative photomicrographs of the immune response stained for Iba1 (microglia), MHCII (reactive microglia), CD4 (T helper cells) and CD8 (cytotoxic T cells) in the SN 5 months after co-injection of different patient-derived homogenates (left) or PMCA-amplified αSYN assemblies (right) with rAAV2/7-αSYN overexpression. Cells immunoreactive for CD4 and CD8 are marked with black arrows. Scale bar Iba1 = 50 µm, MHCII – CD4 = 100 µm. b Table with scoring system for the different immune markers Iba1, MHCII, CD4 and CD8 in the rat SN 5 months after inoculation with patient-derived homogenates (left) or PMCA-amplified αSYN assemblies (right) combined with human αSYN expression. The scoring system ranges from not present (−) to abundantly present (++++). A total of three animals per group was included. The data were presented as a table with scoring system to demonstrate the presence of an inflammatory response in the rat SN

Article Snippet: IHC was performed on free-floating sections using antibodies against tyrosine hydroxylase (TH, rabbit polyclonal, Ab152, 1:5000, Merck Millipore, Massachusetts, US), phosphorylated αSYN (P-S129 αSYN, mouse 11A5, 1:5000, provided by Elan Pharmaceuticals, Inc., Dublin, Ireland and rabbit, Ab51253, 1:5000, Abcam, Cambridge, UK), Iba1 (goat polyclonal, Ab107159, 1:1000, Abcam), MHC Class II (mouse, MCA46G, 1:250, Serotec), CD4 (mouse, MCA55GA, 1:500, Serotec) and CD8 (mouse, MCA48GA, 1:500, Serotec).

Techniques: Derivative Assay, Amplification, Expressing, Staining, Injection, Over Expression