human anti-ifn-γ neutralizing antibodies  (Amgen)

Journal: Scientific Reports

Article Title: Dynamic reciprocal interactions between activated T cells and tumor associated macrophages drive macrophage reprogramming and proinflammatory T cell migration within prostate tumor models

doi: 10.1038/s41598-024-75265-9

Figure Lengend Snippet: IFNG neutralization impacts T cell effect on MDMs. ( A ) Primary M0 MDMs were cultured stepwise in mono-culture, to co-culture with established LNCaP or 22Rv1 tumor cultures; T cells were added for the last 24 h with vehicle control (Ctrl), or IgG control (IgG Iso), or anti-IFNG neutralizing antibody (anti-IFNG). ( B ) IFNG protein concentration (pg/ml) in cell culture supernatants; n = 5; data expressed as mean ± SEM. ( C ) MDM mRNA expression in the six tri-culture cellular and treatment conditions shown as normalized relative quantity (NRQ) as related to house-keeping genes RPLP0 and POLR2A . One-way ANOVA nonparametric with Friedman post-hoc analysis comparing the mean rank of each column with the rank of every other column; n = 5; Data represent mean ± SEM. ( D , F ) Concentrations of secreted cytokines and chemokines in supernatant from the six tri-culture cellular and treatment conditions, expressed as pg/mL. One-way ANOVA nonparametric with Friedman post-hoc analysis comparing the mean rank of each column with the rank of every other column; n = 5; Data represent mean ± SEM. ( E ) T cell migration analysis measured by in-chip confocal microscopy imaging in individual microdevice wells. Data is expressed as each sample’s fold change in migration distance as relative to its corresponding tri-culture vehicle control condition quantified by NIS-Elements. One-way ANOVA nonparametric with Friedman post-hoc analysis comparing the mean ranks of vehicle control IgG control, and vehicle control to Anti-IFNG neutralizing antibody; n = 5; Data represent mean ± SEM.

Article Snippet: For migration experiments, T cells were seeded on Stacks hydrogel wells at a concentration of either 5 × 10 5 or 2.5 × 10 4 cells per ml and allowed to migrate for 24 h. For IFNG neutralization experiments, either 1 μg ml − 1 anti-IFNG neutralizing antibody (R&D Systems by bio-techne, Minneapolis, MN USA Cat# AF-285-NA, RRID: AB_354445) (1:1000), 1 μg ml − 1 IgG control R&D Systems by bio-techne, Minneapolis, MN USA Cat# AB-108-C, RRID: AB_354267) (1:1000), or 0.1% 1xPBS (1:1000) were added to a T cell resuspension media immediately before the addition of the T cells to their culture layer for the 24 h migration period.

Techniques: Neutralization, Cell Culture, Co-Culture Assay, Control, Protein Concentration, Expressing, Migration, Confocal Microscopy, Imaging

Journal: Scientific Reports

Article Title: Dynamic reciprocal interactions between activated T cells and tumor associated macrophages drive macrophage reprogramming and proinflammatory T cell migration within prostate tumor models

doi: 10.1038/s41598-024-75265-9

Figure Lengend Snippet: TAMs retain proinflammatory plasticity in the TME. Dynamic, reciprocal interaction between tumor-polarized macrophages and activated T cells allows paracrine reprogramming of TAM via IFNG secretion.

Article Snippet: For migration experiments, T cells were seeded on Stacks hydrogel wells at a concentration of either 5 × 10 5 or 2.5 × 10 4 cells per ml and allowed to migrate for 24 h. For IFNG neutralization experiments, either 1 μg ml − 1 anti-IFNG neutralizing antibody (R&D Systems by bio-techne, Minneapolis, MN USA Cat# AF-285-NA, RRID: AB_354445) (1:1000), 1 μg ml − 1 IgG control R&D Systems by bio-techne, Minneapolis, MN USA Cat# AB-108-C, RRID: AB_354267) (1:1000), or 0.1% 1xPBS (1:1000) were added to a T cell resuspension media immediately before the addition of the T cells to their culture layer for the 24 h migration period.

Techniques:

Journal:

Article Title: Interferon-? Secretion by t(9;22) Acute Lymphoblastic Leukemia-Derived Dendritic Cells

doi: 10.1016/j.leukres.2010.09.003

Figure Lengend Snippet: IFN-γ secretion was measured from ALL-DC generated from three cell lines and seven patient samples on day 8 of the standard cytokine culture.

Article Snippet: The neutralizing goat anti-human IFN-γ antibody and goat IgG isotype were purchased from Sigma (St. Louis, MO).

Techniques: Generated

Journal:

Article Title: Interferon-? Secretion by t(9;22) Acute Lymphoblastic Leukemia-Derived Dendritic Cells

doi: 10.1016/j.leukres.2010.09.003

Figure Lengend Snippet: (A) IFN-γ secretion was measured by ELISA and (B) immunophenotype was measured by flow cytometry from immature OM9;22 ALL-DC treated with different maturating agent combinations.

Article Snippet: The neutralizing goat anti-human IFN-γ antibody and goat IgG isotype were purchased from Sigma (St. Louis, MO).

Techniques: Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: BMC Cancer

Article Title: Activated CD4 + T cells enhance radiation effect through the cooperation of interferon-γ and TNF-α

doi: 10.1186/1471-2407-10-60

Figure Lengend Snippet: Role of cytokines in the radiosensitization of tumor cells . On day -2, Hela cells were pre-treated for 2 days with IFN-γ; combination of 10 cytokines, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p40, IL-13, IFN-γ, TNF-α and GM-CSF, (Cyt); combination of cytokines without IFN-γ (Cyt-IFN); aCD4S; or combination of cytokines in the presence of neutralizing anti-IFN-γ antibody. On day 1, the cells were treated (or not) with 4 Gy γ-irradiation. Cells were plated in 96-well plate at 5 × 10 3 cells/well in 200 μl CM and cell growth inhibition was determined on day 5 by WST-1 assay. All experiments were performed in triplicate and repeated at least 2 times. Data represent mean values of each triplicate ± S.D.

Article Snippet: For neutralization experiments, monoclonal neutralizing anti-human IFN-γ antibody (eBioscience, CA) was used to block IFN-γ in 1 × 10 5 aCD4S, while mouse IgG1 isotype was used as control.

Techniques: Irradiation, Inhibition, WST-1 Assay

Journal: BMC Cancer

Article Title: Activated CD4 + T cells enhance radiation effect through the cooperation of interferon-γ and TNF-α

doi: 10.1186/1471-2407-10-60

Figure Lengend Snippet: Enhancement of radiosensitizing effect of IFN-γ by TNF-α . On day -2, Hela cells were pre-sensitized for 2 days with IFN-γ, TNF-α, IFN-γ+TNF-α, combination of 10 cytokines (Cyt), or aCD4S, followed (or not) on day 1 with 4 Gy γ-irradiation. Cells were then plated in 96-well plate at 5 × 10 3 cells/well in 200 μl CM and cell growth inhibition was determined on day 5 by WST-1 assay. All experiments were performed in triplicate and repeated at least 2 times. Data represent mean values of each triplicate ± S.D.

Article Snippet: For neutralization experiments, monoclonal neutralizing anti-human IFN-γ antibody (eBioscience, CA) was used to block IFN-γ in 1 × 10 5 aCD4S, while mouse IgG1 isotype was used as control.

Techniques: Irradiation, Inhibition, WST-1 Assay

Journal: BMC Cancer

Article Title: Activated CD4 + T cells enhance radiation effect through the cooperation of interferon-γ and TNF-α

doi: 10.1186/1471-2407-10-60

Figure Lengend Snippet: Effect of aCD4 and cytokine presentization on the redistribution of cell cycle phases in Hela cells . Representative flow cytometric profiles of Hela cells pre-treated for 2 days with IFN-γ, TNF-α, IFN-γ+TNF-α, or aCD4S, followed (or not) with 4 Gy γ-irradiation. The percentage of cells in G0/G1, G2/M or S-phase of the cell cycle is indicated under each curve. These experiments were repeated at least 2 times.

Article Snippet: For neutralization experiments, monoclonal neutralizing anti-human IFN-γ antibody (eBioscience, CA) was used to block IFN-γ in 1 × 10 5 aCD4S, while mouse IgG1 isotype was used as control.

Techniques: Irradiation

Journal: BMC Cancer

Article Title: Activated CD4 + T cells enhance radiation effect through the cooperation of interferon-γ and TNF-α

doi: 10.1186/1471-2407-10-60

Figure Lengend Snippet: General role of aCD4 or IFN-γ/TNF-α combination in the radio-sensitization of cancer cells . Cancer cells from various histology were pre-treated with aCD4S or the combination of 160 U/ml of IFN-γ and 11.2 ng/ml of TNF-α were used to presensitize tumor cells for 3 days, followed by irradiation (4 Gy for a cervical cell line CaSki and prostate cancer cell line DU145, and 8 Gy for a glioma cell line U373). Similar results were seen in two independent experiments. All experiments were performed in triplicate and repeated at least 2 times. Data represent mean values of each triplicate ± S.D. Groups labeled * (p < 0.05) and ** (P < 0.01) were compared to irradiation alone group.

Article Snippet: For neutralization experiments, monoclonal neutralizing anti-human IFN-γ antibody (eBioscience, CA) was used to block IFN-γ in 1 × 10 5 aCD4S, while mouse IgG1 isotype was used as control.

Techniques: Irradiation, Labeling