Anti ifn γ

Manufactured by BioLegend

Sourced in United States, Germany

Anti-IFN-γ is a laboratory reagent used to detect and quantify the presence of interferon gamma (IFN-γ) in biological samples. It functions as a specific antibody that binds to IFN-γ, enabling its identification and measurement.

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Lab products found in correlation

Anti il 4, by BioLegend (62 mentions) Anti cd3, by BioLegend (29 mentions) Anti tnf α, by BioLegend (22 mentions) Anti cd4, by BioLegend (22 mentions) Anti il 17a, by BioLegend (18 mentions) Interleukin 6 (il 6), by BioLegend (17 mentions) Ionomycin, by Merck Group (15 mentions) Tgf β, by BioLegend (12 mentions) Anti il 12, by BioLegend (12 mentions) Anti cd8, by BioLegend (12 mentions) Anti cd28, by BioLegend (11 mentions) Interleukin 6 (il 6), by Thermo Fisher Scientific (10 mentions) Tgf β, by Thermo Fisher Scientific (10 mentions) Anti cd45, by BioLegend (10 mentions) Anti il 10, by BioLegend (9 mentions) Interleukin 2 (il 2), by BioLegend (9 mentions) Anti foxp3, by BioLegend (9 mentions) Golgistop, by BD (9 mentions) Interleukin 4 (il 4), by Thermo Fisher Scientific (9 mentions) Phorbol myristate acetate (pma), by Merck Group (8 mentions)

Spelling variants of this product

IFN-γ (422 citations) Anti-IFN-γ-APC (38 citations) Anti-IFN-γ antibody (22 citations) Anti-IFN-γ–APC–Cy7 (7 citations) Anti-IFN-γ-PE (clone 4S.B3; (3 citations) PerCP anti-human IFN-γ (2 citations) Anti-Mouse-IFN-γ-FITC (XMG 1.2) (2 citations) Anti-IFN-γ-BV605 (2 citations) Anti–IFN-γ and anti-TNFα (2 citations) FITC-labeled anti-IFN-γ (2 citations)

149 protocols using anti ifn γ

Polarization of Naive CD4+ T Cells

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Naive CD4⁺ T cells were enriched from PBMCs by negative magnetic separation using MojoSort Human CD4 Naive T cell isolation Kit (BioLegend). The purity of isolated cells was checked by flow cytometry (> 95%). 2.5 × 10⁵ naive cells were cultured in flat-bottom 96-well plates with CD3/CD28 Dynabeads in RPMI 1640 supplemented with 15% FCS, 10 mM HEPES (Sigma), 100 U/ml penicillin (Sigma), and 200 mM L-glutamine (Sigma) for 4 days under the following conditions: Th0 (anti-IL-4 (5 μg/ml), anti-IFNγ (5 μg/ml)); Th1 (anti-IL-4 (5 μg/ml), IL-12 (10 ng/ml)); Th2 (IL-4 (10 ng/ml), anti-IFNγ (5 μg/ml)); Th17 (IL-6 (20 ng/ml), TGFβ (1 ng/ml), anti-IFNγ (5 μg/ml), anti-IL-4 (5 μg/ml)) (all recombinant proteins and antibodies were from BioLegend). On day 4, cells were stimulated with PMA (50 ng/ml) and ionomycin (375 ng/ml) for 5 h. Golgi Plug was added into the culture (1:1000 dilution) 1 h after stimulation. The levels of IFNγ, IL-4, and IL-17A were measured by flow cytometry with intracellular staining protocol.

Aba Ü., Maslak İ.C., İpşir C., Pehlivan D., Warnock N.I., Tumes D.J., Cildir G, & Erman B. (2024). A Novel Homozygous Germline Mutation in Transferrin Receptor 1 (TfR1) Leads to Combined Immunodeficiency and Provides New Insights into Iron-Immunity Axis. Journal of Clinical Immunology, 44(2), 55.

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In Vitro T Cell Differentiation

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OT-2 T cells and OVA-loaded APCs were prepared as above. Naive OT-2 T cells (5×10⁴) and purified OVA-loaded antigen-presenting cells (1×10⁴) were co-cultured in 96-well round-bottom tissue culture plates under the control (medium alone) or Th polarizing conditions (23 (link)) as follows: Th1 with 10 ng/ml IL-12 (BioLegend) and 5 μg/ml anti-IL-4 (BioLegend); Th2 with 10 ng/ml IL-2 (BioLegend), 10 ng/ml IL-4 (BioLegend), and 5 μg/ml anti-IFN-γ (BioLegend); Th17: 2 ng/ml TGF-β (Peprotech, Rocky Hill, NJ), 10 ng/ml IL-1β (BioLegend), 20 ng/ml IL-6 (BioLegend), 5 μg/ml anti-IFN-γ, and 5 μg/ml anti-IL-4. Intracellular staining of Th cytokines IFN-γ (Th1), IL-4 (Th2), and IL-17A (Th17) were performed by flow cytometry following stimulation with PMA, ionomycin, and brefeldin A at 3 to 5 days after culture.

Ryu S.H., Shin H.S., Eum H.H., Park J.S., Choi W., Na H.Y., In H., Kim T.G., Park S., Hwang S., Sohn M., Kim E.D., Seo K.Y., Lee H.O., Lee M.G., Chu M.K, & Park C.G. (2022). Granulocyte Macrophage-Colony Stimulating Factor Produces a Splenic Subset of Monocyte-Derived Dendritic Cells That Efficiently Polarize T Helper Type 2 Cells in Response to Blood-Borne Antigen. Frontiers in Immunology, 12, 767037.

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Characterizing Tumor-Infiltrating CD8+ T Cells

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CD8 + T cells from tumor-infiltrating lymphocytes were isolated using flow cytometry-assisted cell sorting. The purity of the sorted cells was >95 %. To detect cytokine production, lymphocytes were stimulated for 2 h in the presence of a cell-stimulating mixture (eBioscience, USA, 00-4975-93) and then stained with the surface markers anti-LAG-3 (BioLegend, USA, Cat. No. 125214) and anti-PD-1 (BioLegend, Cat. No. 135206), and CTLA4 (BioLegend, Cat. No. 106308) at a 1:100 dilution in PBS containing 1 % FBS. Following a 30 min incubation on ice, a cell fixation/permeabilization kit (BD Biosciences, USA, Cat. No. 554714) and a transcription factor fixation/permeabilization buffer (BioLegend, Cat. No. 424401) were employed for intracellular cytokine and nuclear transcription factor labeling, respectively. Anti-IFN-γ (BioLegend, USA, Cat. No. 505810), anti-TNF-α (BioLegend, Cat. No. 506306), and anti-TOX (Miltenyi Biotec, Germany, Cat. No. 130-120-337) were diluted 1:50 in permeabilization buffer and incubated overnight at 4 °C in the presence of Anti-IFN-γ and anti-TNF-α(BioLegend, USA). FlowJo software (Tree Star, USA) was used to analyze FACSCelesta (BD, USA) flow cytometry data. All analyses were performed using GraphPad Prism 8 (GraphPad Software, USA) and t-tests. P < 0.05 was deemed statistically significant.

Wu D., Wang G., Wen S., Liu X, & He Q. (2024). ARID5A stabilizes Indoleamine 2,3-dioxygenase expression and enhances CAR T cell exhaustion in colorectal cancer. Translational Oncology, 42, 101900.

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Multiparametric Flow Cytometry of Immune Cells

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Different panels of antibody cocktails were generated from anti‐CD45 (BioLegend, 103116, 30‐F11), anti‐CD3 (BioLegend, 100218, 17A2), anti‐CD4 (BioLegend, 100421, GK1x.5), anti‐CD8a (BioLegend, 100711, 53‐6.7), anti‐NKp46 (BioLegend, 137606, 29A1.4), anti‐CD11c (BioLegend, 117307, N418), anti‐CD25 (Biolegend, 101908, 3C7), anti‐FOXP3 (Biolegend, 126404, MF‐14), anti‐granzyme B (BioLegend, 372208, QA16A02), anti‐IFNγ (BioLegend, 505849, XMG1.2), anti‐IFNγ (BioLegend, 505806, XMG1.2, anti‐CD62L (BioLegend, 104432, MEL‐14) and anti‐CD44 (BD Biosciences, 560568, IM7) antibodies, and the AmCyan Live/Dead Cell Staining Kit (Thermo Fisher Scientific). All antibodies were diluted at optimized dilutions before being used.
Female 7–8 weeks old C57BL/6 mice (n = 4–5 per group) were intravenously administered with the equivalent dose (3.8 × 10¹⁰ vg/mouse) of either free AAV‐fLuc or RBC‐AAV‐fLuc. Mice were then euthanized on day 31, and spleen and lungs were collected. Organ dissociation kits (Miltenyi Biotec) were used per manufacturer's instruction to generate a single‐cell suspension from excised organs, and cells were stained with the antibodies listed above and analyzed using flow cytometry (BD LSR II Analyser). Flow cytometry data analyses were performed using FlowJo 10 software.

Zhao Z., Kim J., Suja V.C., Kapate N., Gao Y., Guo J., Muzykantov V.R, & Mitragotri S. (2022). Red Blood Cell Anchoring Enables Targeted Transduction and Re‐Administration of AAV‐Mediated Gene Therapy. Advanced Science, 9(24), 2201293.

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Activation of Jurkat and Naive CD4+ T cells

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Jurkat and naïve CD4⁺ T cells were activated with plate bounded anti-CD3 and CD28 (anti-mouse CD3, Invitrogen, anti-mouse CD28 and anti-human CD3/28, BioLegend, 2.5 μg/ml each) for the indicated time. Other than anti-CD3 and CD28, conditions for Th0 and Tfh-like cells: Th0 (anti-IL4 10 μg/ml, anti-IFNγ 10 μg/m, BioLegend); Tfh-like (anti-IL4 10 μg/ml, anti-IFNγ 10 μg/ml, anti-IL12 10 μg/ml, anti-TGFβ 10 μg/ml, BioLegend; IL6 100 ng/ml and IL21 50 ng/ml, PeproTech).
For the T cell coculture regents, rapamycin was purchased from Cell Signaling Technology. 4EGI-1 and Cycloheximide (CHX) were purchased from MedChemExpress. Cantharidin (CAN), okadaic acid (OA), and MG132 were purchased from Selleckchem. Sphingomyelinase (SMase) was purchased from Merck.

Sanna M.G., Duckett C.S., Richter B.W., Thompson C.B, & Ulevitch R.J. (1998). Selective activation of JNK1 is necessary for the anti-apoptotic activity of hILP. Proceedings of the National Academy of Sciences of the United States of America, 95(11), 6015-6020.

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Naïve CD4+ T Cell Polarization

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Naive CD4⁺ T cells were purified by mouse CD4⁺CD62L⁺ T Cell Isolation Kit II (Miltenyi Biotec). Purified naive T cells were stimulated with plate-bound goat anti-hamster antibodies, soluble anti-CD3 (0.25 μg ml⁻¹,145-2C11; Biolegend) and anti-CD28 (0.5 μg ml⁻¹, 37.51; Biolegend) for Th0-non-polarized condition culture. In addition to Th0-non-polarized condition, following stimulation was used for each polarized condition: IL-12 (20 ng ml⁻¹; R&D Systems) and anti-IL-4 (10 μg ml⁻¹, C17.8; Biolegend) for Th1; IL-4 (100 ng ml⁻¹; R&D Systems), anti-IL-12 (10 μg ml⁻¹; Biolegend) and anti-IFNγ (10 μg ml⁻¹, XMG1.2; Biolegend) for Th2; IL-6 (3 ng ml⁻¹ or indicated concentration; R&D Systems), TGF-β1 (0.3 ng ml⁻¹; R&D Systems), anti-IL-4 (10 μg ml⁻¹, C17.8; Biolegend) and anti-IFNγ (10 μg ml⁻¹; XMG1.2; Biolegend) for Th17; and IL-2 (20 ng ml⁻¹; R&D Systems), TGF-β1 (3 ng ml⁻¹; R&D Systems), anti-IL-4 (10 μg ml⁻¹, C17.8; Biolegend) and anti-IFNγ (10 μg ml⁻¹, XMG1.2; Biolegend) for Tregs. For signal transduction studies, STA21 (STAT3 inhibitor; Santa Cruz Biotechnology) and SMAD3 inhibitor (SIS3; Santa Cruz Biotechnology) were added to cultures on day 0.

Yoshida N., Comte D., Mizui M., Otomo K., Rosetti F., Mayadas T.N., Crispín J.C., Bradley S.J., Koga T., Kono M., Karampetsou M.P., Kyttaris V.C., Tenbrock K, & Tsokos G.C. (2016). ICER is requisite for Th17 differentiation. Nature Communications, 7, 12993.

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Naïve T-cell Polarization Assay

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Spleen from five naïve C57BL/6 mice were harvested and a single cell suspension was prepared followed by RBC lysis. Naïve T cells were negatively selected using EasySep™ Mouse T-cell Isolation Kit (Stemcell Technologies, Vancouver Canada) as per manufacturer’s instructions. Naïve T cells were then polarized in to different subsets as described previously (Noubade et al. 2011 (link); Noubade et al. 2007 (link)). Briefly, 1×10⁵ T cells were stimulated for 72h in complete RPMI media (Genesee, San Diego, CA) with plate bound anti-CD3 (5μg/ml) and soluble CD28 antibodies (2μg/ml) (Biolegend) for the T_h0 condition. In addition, the following cytokines and antibodies were added to the T_h0 conditions for polarization: T_h1 (10μg/ml anti-IL-4 (BioLegend), 4ng/ml IL-12 (Peprotech)), T_h2 (10μg/ml anti-IFN-γ (Biolegend), 30ng/ml IL-4 (Peprotech)), T_h17 (30ng/ml IL-6, 1ng/ml TGF-β (Biolegend), 10μg/ml anti-IFN-γ, 10μg/ml anti-IL-4) and T_reg (2ng/ml TGF-β). Cultures were harvested for RNA and supernatants.

Ginwala R., Bhavsar R., Moore P., Bernui M., Singh N., Bearoff F., Nagarkatti M., Khan Z.K, & Jain P. (2020). Apigenin modulates dendritic cell activities and curbs inflammation via RelB inhibition in the context of neuroinflammatory diseases. Journal of neuroimmune pharmacology : the official journal of the Society on NeuroImmune Pharmacology, 16(2), 403-424.

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Isolation and Differentiation of Human CD4+ T Cells

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Human peripheral blood was obtained from healthy volunteers who are not taking any medications. PBMCs were prepared by using lymphocyte separation medium (MP Biomedicals), according to the manufacturer’s instructions. Subsequently, total CD4⁺ T cells were isolated by means of negative selection with CD14 microbeads (Miltenyi Biotec), followed by positive selection with CD4 microbeads (Miltenyi Biotec). Naive and effector memory CD4⁺ T cells were further sorted as CD3⁺CD4⁺ CD45RA⁺CD45RO⁻CD25⁻ and CD3⁺CD4⁺CD45RA⁻CD45RO⁺ cells, respectively, by using the FACSAria III cell sorter (BD Biosciences). For naive T-cell differentiation, naive T cells undergoing fluorescence-activated cell sorting (FACS) were stimulated with plate-coated anti-CD3 (10 μg/mL, OKT3; BioLegend) and soluble anti-CD28 (2 μg/mL, CD28.2; BioLegend) for 4 or 7 days. For T_H2 polarization, IL-2 (10 ng/mL), IL-4 (10 ng/mL), and anti–IFN-γ (5 μg/mL, B27; BioLegend) were added. For T_H17 polarization, IL-2 (10 ng/mL), IL-6 (20 ng/mL), TGF-β (10 ng/mL), IL-23 (20 ng/mL), IL-21 (20 ng/mL), IL-1β (20 ng/mL), anti–IFN-γ (5 μg/mL), and anti–IL-4 (5 μg/mL, MP4-25D2; BioLegend) were added. TGF-β was from PeproTech, and the rest of the cytokines were from eBioscience. Effector memory CD4⁺ T cells were stimulated with plate-coated anti-CD3 (10 μg/mL) for 48 hours in the presence of UA or DMSO.

Na H., Lim H., Choi G., Kim B.K., Kim S.H., Chang Y.S., Nurieva R., Dong C., Chang S.H, & Chung Y. (2017). Concomitant suppression of TH2 and TH17 cell responses in allergic asthma by targeting retinoic acid receptor–related orphan receptor γt. The Journal of allergy and clinical immunology, 141(6), 2061-2073.e5.

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Naïve CD4 T Cell Differentiation

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Naïve CD4 T cells were isolated by sorting CD4⁺CD25^-CD44^lowCD62L⁺ T cells by FACS. Purified CD4⁺ T cells were stimulated with plate-bound anti-CD3 at 5ug/ml and anti-CD28 at 2ug/ml antibodies in the presence or absence of IL12 (100ng/ml) or IFNγ at different concentrations for the indicated times. Th differentiation was carried out by incubation of naïve CD4 T cells with 5ug/ml anti-CD3 and 2ug/ml anti-CD28 together with: for Th0, 20ng/ml IL2 (R&D system 402-ML-020); for Th1, 20ng/ml IL2, 20ng/ml IL12 (R&D system 419-ML-010), 10ug/ml anti-IL4 (Biolegend 504102); for Th2 20ng/ml IL2, 100ng/ml IL4 (Biolegend 574302), 10ug/ml anti-IL12 (Biolegend 505303), 10ug/ml anti-IFNγ (Biolegend 505702); for Th17 2ng/ml TGFβ (R&D system 240-B-002), 100ng/ml IL6 (Biolegend 575702), 10ug/ml anti-IL4, 10ug/ml anti-IFNγ, for 5 days. For analysis of cytokine producing cells the cultures were stimulated for three hours with 100ng/ml PMA and 200ng/ml Ionomycin and Golgistop (BD) and analyzed by intracellular cytokine staining. Egr2 and T-bet staining was analyzed using the Foxp3 staining kit (E-bioscience).

Singh R., Miao T., Symonds A.L., Omodho B., Li S, & Wang P. (2017). Egr2 and 3 Inhibit T-bet–Mediated IFN-γ Production in T Cells. The Journal of Immunology Author Choice, 198(11), 4394-4402.

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In Vitro Th17 and iTreg Polarization

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Teff cells (3 × 10⁵ cells/well) were cultured in complete RPMI-1640 medium and stimulated with Dynabeads^® Mouse T-Activator CD3/CD28 (8 μl/well) for 72 h along with cytokines and neutralizing antibodies for the desired polarization as follows: human-IL-6 (100 ng/ml; PeproTech. INC.), mouse-IL-23 (10 ng/ml; Biolegend), human-TGF-β (5 ng/ml), anti-IL4 (10 μg/ml; Biolegend) and anti-IFN-γ (10 μg/ml; Biolegend) for Th17; mouse IL-2 (10 ng/ml), human-TGF-β (2 ng/ml), anti-IL4 (10 μg/ml) and anti-IFN-γ (10 μg/ml) for inducible Treg (iTreg) cell polarization. Cells were restimulated with PMA (50 ng/ml), ionomycin (1 μg/ml) and brefeldin A (1 μg/ml) for 6 h. Cells were harvested and stained with anti-IL-17 or anti-Foxp3 antibodies and analyzed by flow cytometry.

Kim D., Kim J.Y, & Jun H.S. (2016). Smad4 in T cells plays a protective role in the development of autoimmune Sjögren's syndrome in the nonobese diabetic mouse. Oncotarget, 7(49), 80298-80312.

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