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

Article Title: Adoptive transfer of Tc1 or Tc17 cells elicits anti-tumor immunity against established melanoma through distinct mechanisms 1

doi: 10.4049/jimmunol.1201989

Figure Lengend Snippet: Generation and Tc1 and Tc17 Responses of antigen-specific to tumor antigen stimulation ex vivo. (A) Tc1 and Tc17 cells were generated as described in “Materials and Methods”, and their phenotype in terms of IFNγ and IL-17 production was shown on gated Pmel-1 (Thy1.1+) cells. (B) In a separate experiment, pmel-1 Tc1 and Tc17 cells were labeled with CSFE and transferred into B16 tumor-bearing B6 mice (n = 4). Five days after cell transfer, mice were euthanized and their splenocytes were stained for CD8 and Thy1.1. The CSFE profile is shown on gated live CD8+Thy1.1+ cells for one of four representative mice. (C) A set of B6 mice (n = 3) were inoculated i.v. with luciferase transduced B16 F10 cells and received Tc1 or Tc17 cells combined with BMT. Mice were euthanized on day 14 after treatment. Splenocytes were isolated and processed for cytokine-release ELISPOT assays. Gp100-specific IFNγ- and IL-17-producing cells were detected after co-culture with gp100-pulsed and unpulsed EL4 for 20 hours. Experiments were repeated 2 times with similar results. (D) The graph shows the average number of IFNγ- and IL-17-producing spots from triplicate wells with 1SD as error bars.

Article Snippet: In brief, splenocytes from Pmel-1 transgenic mice were stimulated with the hgp25–33 peptide (1 μg/ml), recombinant mouse IL-12 (10 ng/ml; R&D Systems) and recombinant mouse IFNγ (1000 U/ml R&D Systems).

Techniques: Ex Vivo, Generated, Labeling, Staining, Luciferase, Isolation, Enzyme-linked Immunospot, Co-Culture Assay

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

Article Title: Adoptive transfer of Tc1 or Tc17 cells elicits anti-tumor immunity against established melanoma through distinct mechanisms 1

doi: 10.4049/jimmunol.1201989

Figure Lengend Snippet: Requirement of IFNγR on tumor cells for Tc1- or Tc17-mediated anti-tumor responses. (A) B6 mice were inoculated i.v. with IFNγRDN B16 tumor cells. gp100 specific Tc1 and Tc17 cells were administered 7 days after tumor implantation. Overall survival rate was observed. (B) Ag-specific IL-17 cytokine-release was analyzed by an ELISPOT assay using splenocytes from IFNγRDN B16 tumor bearing mice. ELISPOT was performed using several stimulator cells: IFNγ-treated (100 U/mL, 20 hours), and non-treated and IFNγ-treated (100 U/mL, 20 hours) B16, and non-treated B16 IFNγRDN B16 tumor. (C) Ag-specific IFNγ and cytokine-release was analyzed by ELISPOT assays using splenocytes from IFNγRDN B16 tumor bearing mice. ELISPOT was performed using several stimulator cells: IFNγ-treated (100 U/mL, 20 hours), and non-treated B16 and non-treated B16 IFNγRDN B16 tumor.

Article Snippet: In brief, splenocytes from Pmel-1 transgenic mice were stimulated with the hgp25–33 peptide (1 μg/ml), recombinant mouse IL-12 (10 ng/ml; R&D Systems) and recombinant mouse IFNγ (1000 U/ml R&D Systems).

Techniques: Tumor Implantation, Enzyme-linked Immunospot

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

Article Title: Adoptive transfer of Tc1 or Tc17 cells elicits anti-tumor immunity against established melanoma through distinct mechanisms 1

doi: 10.4049/jimmunol.1201989

Figure Lengend Snippet: Role of IFN-γ and IL-17 in Tc17-mediated anti-tumor effect. WT C57/B6 mice (n = 4 per group) were given B16 (A) and IFNγRDN B16 tumor (B) i.v. by tail vein. Seven days later, tumor-bearing mice were adoptively transferred with Tc1 and Tc17 cells in the combination of BM. Anti-IFNγ mAb and anti-IL-17 mAb was administered intraperitoneally on days 0 and continuously twice a week for 4 weeks total. Overall survival was monitored. Corresponding groups of untreated tumor-bearing mice served as controls.

Article Snippet: In brief, splenocytes from Pmel-1 transgenic mice were stimulated with the hgp25–33 peptide (1 μg/ml), recombinant mouse IL-12 (10 ng/ml; R&D Systems) and recombinant mouse IFNγ (1000 U/ml R&D Systems).

Techniques:

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

Article Title: Adoptive transfer of Tc1 or Tc17 cells elicits anti-tumor immunity against established melanoma through distinct mechanisms 1

doi: 10.4049/jimmunol.1201989

Figure Lengend Snippet: Activation and expansion of antigen-specific Tc1 and Tc17 cells in vivo. Normal B6 mice with established WT or IFNγRDN B16 tumor received Tc1 or Tc17 cells in combination with BMT. Four mice were euthanized on day 14 and 28 after ACT. (A) Leukocytes isolated from spleen or lung on day 14 were stained with anti-Thy 1.1, anti-CD8, anti-IFNγ, anti-IL-17 and TNFα. Intracellular cytokine expression was shown on gated Thy 1.1+ cells. Flow plots display 1 of 3-4 mice in each group. (B) Percentages of IFNγ+, /IL-17+, IFNγ+/IL-17+and TNFα+ on gated Thy 1.1+ cells are shown from 3-4 mice per group on day 14 in spleen and lung. (C) Absolute numbers of adoptive transferred Tc1 or Tc17 cells are shown from 3-4 mice per group in spleen and lung on day 14 and 28. The data represent 1 of 3 experiments using similar settings. * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: In brief, splenocytes from Pmel-1 transgenic mice were stimulated with the hgp25–33 peptide (1 μg/ml), recombinant mouse IL-12 (10 ng/ml; R&D Systems) and recombinant mouse IFNγ (1000 U/ml R&D Systems).

Techniques: Activation Assay, In Vivo, Isolation, Staining, Expressing

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

Article Title: Adoptive transfer of Tc1 or Tc17 cells elicits anti-tumor immunity against established melanoma through distinct mechanisms 1

doi: 10.4049/jimmunol.1201989

Figure Lengend Snippet: Phenotype of Tc1 and Tc17 cells after ACT

Article Snippet: In brief, splenocytes from Pmel-1 transgenic mice were stimulated with the hgp25–33 peptide (1 μg/ml), recombinant mouse IL-12 (10 ng/ml; R&D Systems) and recombinant mouse IFNγ (1000 U/ml R&D Systems).

Techniques:

Journal: Vaccines

Article Title: Development of a T Cell-Based COVID-19 Vaccine Using a Live Attenuated Influenza Vaccine Viral Vector.

doi: 10.3390/vaccines10071142

Figure Lengend Snippet: Figure 9. The levels of IFNγ-secreting cells in the splenocytes of immunized Syrian hamsters (n = 4) on day 5 after challenge with SARS-CoV-2 Wuhan (A) or Delta (B) variants. Isolated splenocytes were stimulated in vitro either with live SARS-CoV-2 or with PepTivator, followed by quantification of IFNγ-secreting cells by Hamster IFN-γ ELISpot Plus kit. Data were analyzed by two-way ANOVA with Tukey’s post-hoc multiple analyses test. *—p < 0.05; **—p < 0.01.

Article Snippet: Assessment of Cellular Responses by ELISpot Assay Splenocytes isolated from mice and Syrian hamsters were processed identically, except that different ELISpot kits were used to detect IFNγ production: Mouse IFN-γ Single-Color ELISPOT (Cellular Technology Ltd., Shaker Heights, OH, USA) and Hamster IFN-γ ELISpot Plus kit (Mabtech, Nacka Strand, Sweden) using the instructions of the manufacturers.

Techniques: Isolation, In Vitro, Enzyme-linked Immunospot

Journal: Microbial biotechnology

Article Title: Rectal single dose immunization of mice with Escherichia coli O157:H7 bacterial ghosts induces efficient humoral and cellular immune responses and protects against the lethal heterologous challenge

doi: 10.1111/j.1751-7915.2011.00316.x

Figure Lengend Snippet: Cellular immune responses stimulated in mice after the immunizations with EHEC O157:H7 (N°CIP 105282) BGs. Mice were immunized as described in Fig. 1 . Spleen cells (1 × 10 5 per well) were incubated for 16 h in the presence of EHEC O157:H7 BGs (specific Ag); measles inactivated Ag (strain Edmonston, non‐specific Ag) or as a negative control spleen cells were left without stimulation. The numbers of IFN‐γ‐producing cells were determined by ELISPOT assays. Rectal immunization with EHEC O157:H7 BGs effectively increased the number of IFN‐γ‐producing spleen cells after recognition of specific Ag (A). The stimulatory effect of the boost immunization was detected on day 42 (14 days after the second immunization) and remained significant until the end of experiment (B). Data represent the mean of three independent experiments ± SD (three mice per time point). P ‐values < 0.05 were considered significant and are indicated with asterisks (* P < 0.05; ** P < 0.01; *** P < 0.001). a, day of the first immunization in all groups; b, day of the second immunization in groups B1 and D1; c, day of the challenge.

Article Snippet: The number of IFN‐γ‐producing cells was determined by Mouse IFN‐γ ELISpot Kit (R&D Systems, Inc, Minneapolis, USA) following the manufacturer's instructions.

Techniques: Incubation, Negative Control, Enzyme-linked Immunospot

Journal: Drug Design, Development and Therapy

Article Title: Preclinical Assessment of HLA-A*02:01-Restricted PSMA and STEAP1 Epitopes for Peptide-Based Immunotherapy in Prostate Cancer

doi: 10.2147/DDDT.S576346

Figure Lengend Snippet: IFN-γ ELISpot analysis of peptide-specific T-cell responses in HLA-A*02:01 transgenic mice. Splenocytes were harvested ten days after booster immunization and stimulated ex vivo with the indicated peptides. Bars represent mean spot-forming units (SFUs) per 10 5 splenocytes (n = 6 mice per group). Error bars indicate standard error of the mean (SEM). Statistical significance was assessed using one-way ANOVA followed by Tukey’s multiple comparison test. (*** p < 0.001).

Article Snippet: Splenocytes (2 × 10 5 cells/well) were seeded into anti-mouse IFN-γ–precoated ELISpot plates (Mabtech, Sweden) and stimulated with 10 μg/mL of the respective synthetic peptide for 48 hours at 37 °C in a 5% CO 2 incubator.

Techniques: Enzyme-linked Immunospot, Transgenic Assay, Ex Vivo, Comparison

Journal: Drug Design, Development and Therapy

Article Title: Preclinical Assessment of HLA-A*02:01-Restricted PSMA and STEAP1 Epitopes for Peptide-Based Immunotherapy in Prostate Cancer

doi: 10.2147/DDDT.S576346

Figure Lengend Snippet: Integrative heatmap summarizing immunogenic profiling across five criteria—docking score, ELISA binding (OD), IFN-γ ELISpot responses, CD8⁺IFN-γ⁺ T cell percentages, and TNF-α co-expression for each predicted epitope. Color intensity reflects relative performance. ALFDIESKV and FLYTLLREV demonstrate consistently high scores across all parameters.

Article Snippet: Splenocytes (2 × 10 5 cells/well) were seeded into anti-mouse IFN-γ–precoated ELISpot plates (Mabtech, Sweden) and stimulated with 10 μg/mL of the respective synthetic peptide for 48 hours at 37 °C in a 5% CO 2 incubator.

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, Enzyme-linked Immunospot, Expressing

Journal: Frontiers in Immunology

Article Title: Identification of a DRB3*011:01-restricted CD4 + T cell response against bovine respiratory syncytial virus fusion protein

doi: 10.3389/fimmu.2023.1040075

Figure Lengend Snippet: Stimulation of CD4 + T cells with synthetic, overlapping peptides from the BRSV-375 F protein. CD4 + T cells from dairy cows were positively selected and cultured with autologous antigen presenting cells loaded with synthetic peptide. T cell activation was assessed via IFN-γ secretion using ELISpot assays. Only cells from DRB3*011:01 positive animals responded to stimulation with five out of 88 peptides tested. T cell responses from animals 1141 (A) , 1310 (B) , 2938 (C) , and 2942 (D) displayed a measurable T cell response. Peptides sharing N-terminal amino acid with BRSV-375 F protein residue 253 generated a slight response while those with N-terminal amino acids matching residues 257-260 displayed a larger response. T cells were stimulated with PHA at a concentration of 5μg/mL as a positive control, and cRPMI media as a negative control. Data presented as mean ± SEM, p < 0.05 was considered significant and indicated by asterisk.

Article Snippet: The day prior to, 96-well multiscreen filter plates (MilliporeSigma, Burlington, MA, USA) were coated with mouse anti-bovine IFN-γ antibody (Bio-Rad, Hercules, CA, USA) at 4°C overnight.

Techniques: Cell Culture, Activation Assay, Enzyme-linked Immunospot, Generated, Concentration Assay, Positive Control, Negative Control

Journal: Biomarker Research

Article Title: Computation strategies and clinical applications in neoantigen discovery towards precision cancer immunotherapy

doi: 10.1186/s40364-025-00808-9

Figure Lengend Snippet: A representative discovery of neoantigen prediction. Collecting tumor tissues and normal tissues such as para-carcinomatous tissues or peripheral blood mononuclear cells (PBMC) for sequencing. Subsequently, HLA typing is conducted to ascertain accurate HLA subtypes. Further, to generate candidate neoantigens employing Peptide-HLA-TCR binding prediction algorithms. Finally, the immunogenicity of candidate neoantigens is validated through enzyme-linked ImmunoSpot (ELISpot) assays or mouse animal experiments. The validated neoantigens are subsequently utilized for immunotherapy

Article Snippet: Finally, the immunogenicity of candidate neoantigens is validated through enzyme-linked ImmunoSpot (ELISpot) assays or mouse animal experiments.

Techniques: Sequencing, Immunopeptidomics, Binding Assay, Enzyme-linked Immunospot

Journal: Cancer cell

Article Title: MACROPHAGE POLARIZATION CONTRIBUTES TO GLIOBLASTOMA ERADICATION BY COMBINATION IMMUNOVIROTHERAPY AND IMMUNE CHECKPOINT BLOCKADE

doi: 10.1016/j.ccell.2017.07.006

Figure Lengend Snippet: A. 005 GSCs cultured for 24 hr with or without murine IFNγ (mIFNγ: 0 or 3 ng/ml), stained for PD-L1, and analyzed by flow cytometry. B. 005 GSCs infected with G47Δ-E or G47Δ-mIL12 at MOI=1 for 24 hr, stained for PD-L1, and analyzed by flow cytometry. C. Human primary (left) and recurrent (right) GSCs cultured for 24 hr, stained for PD-L1 (cyan), and analyzed by flow cytometry. Percent of PD-L1+ cells indicated in upper right. See also Figure S1.

Article Snippet: Mouse IFNγ ELISPOT assay Mouse IFNγ ELISPOT assay was performed according to the manufacturer instructions (R&D Systems) with minor modifications.

Techniques: Cell Culture, Staining, Flow Cytometry, Infection