Pe cy7 conjugated anti ifn γ

Manufactured by BD

Sourced in United States

PE-Cy7-conjugated anti-IFN-γ is a flow cytometry reagent that consists of a phycoerythrin-cyanine 7 (PE-Cy7) fluorescent label conjugated to an antibody specific for interferon-gamma (IFN-γ). This reagent can be used to detect and quantify IFN-γ-producing cells in various sample types.

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Close spelling variants of this product

IFN-γ (389 citations) Anti-IFN-γ (150 citations) Anti-IFN-γ-PE (76 citations) IFN-γ-PE (64 citations) IFN-γ-PE-Cy7 (42 citations) Anti-IFN-γ PE-Cy7 (25 citations) PE-conjugated anti-IFN-γ (11 citations)

5 protocols using pe cy7 conjugated anti ifn γ

Multiparametric Flow Cytometry of T-cell Subsets

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The stimulated PBMCs were harvested and stained with allophycocyanin (APC)-conjugated anti-CD4 and (APC-H7)-conjugated anti-CD3, fixed and permeabilized with the permeabilization/fixation solution (eBiosciences, San Diego, USA), followed by staining with PE-conjugated anti-IL-22 (R&D Systems, Minneapolis, MN, USA), FITC-conjugated anti-IL-17 (BD Pharmingen, San Diego, CA, USA), and PE-Cy7-conjugated anti-IFN-γ (BD Pharmingen, San Diego, CA, USA). The frequency of CD4⁺IFN-γ^–IL17A^–IL-22⁺ (Th22), CD4⁺IFN-γ^–IL17A⁺IL-22⁻ (Th17), CD4⁺IFN-γ⁻IL17A⁺IL-22⁺(Th17/Th22) and CD4⁺IFN-γ⁺ (Th1) T cells in the samples was determined by flow cytometry analysis on a BD FACSAria II (Becton Dickinson) using FlowJo 7.6.2 software.

Guo H., Peng D., Yang X.G., Wang Y., Xu B.C., Ni J.S., Meng W, & Jiang Y.F. (2014). A Higher Frequency of Circulating IL-22+CD4+ T Cells in Chinese Patients with Newly Diagnosed Hashimoto’s Thyroiditis. PLoS ONE, 9(1), e84545.

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Th1 Cell Immunophenotyping and Cytokine Analysis

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The PBMCs were harvested and stained with allophycocyanin- (APC-) H7 mouse anti-human CD4 and PerCP-Cy™5.5 mouse anti-human CD3 (BD Pharmingen, San Diego, CA, USA) for 30 min. After staining, the cells were fixed and permeabilized using the permeabilization/fixation solution kit (eBioscience, San Diego, USA) for 30 min, followed by staining with PE-Cy7-conjugated anti-IFN-γ (BD Biosciences, San Diego, CA, USA) for 30 min. The frequency of CD3⁺CD4⁺IFN-γ⁺ (Th1) T cells was determined by flow cytometry analysis using the BD FACSAria II (Becton Dickinson, Washington, DC, USA) and analyzed with the FlowJo7.6.2 software (FlowJo LLC, Ashland, OR, USA). IFN-γ and TNF-α plasma levels were determined by cytometric bead array (CBA) analysis. The serum concentrations of cytokines were quantified using the CellQuest Pro and CBA software (Becton Dickinson, Washington, DC, USA) on a FACSCalibur Flow Cytometer (Becton Dickinson).

Xu G., Zhang P., Dang R., Jiang Y., Wang F., Wang B, & Yang M. (2019). Dynamic Changes of Th1 Cytokines and the Clinical Significance of the IFN-γ/TNF-α Ratio in Acute Brucellosis. Mediators of Inflammation, 2019, 5869257.

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Stimulation and Intracellular Cytokine Staining of PBMCs

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PBMCs were suspended in RPMI media (Invitrogen) and rested for 2 hours. One million cells were stimulated in vitro with phorbol 12-myristate 13-acetate and ionomycin or with overlapping peptides from SIV p27 protein for 9 hours at 37°C as previously described (58 (link)). After incubation, cells were washed and immunostained with Alexa Fluor 700–conjugated anti-CD3 (BD Biosciences, SP-234), BV650-conjugated anti-CD4 (BD Biosciences, L200), and PerCyP-Cy5.5–conjugated anti-CD8 (Biolegend, OKT4). Cells were washed, permeabilized using a Cytofix/Cytoperm Kit (BD Biosciences), and intracellularly immunostained with PE-Cy7–conjugated anti–IFN-γ (BD Biosciences, B27) and Alexa Fluor 488–conjugated anti–TNF-α (Invitrogen, Mab11).

Weber M.G., Walters-Laird C.J., Kol A., Santos Rocha C., Hirao L.A., Mende A., Balan B., Arredondo J., Elizaldi S.R., Iyer S.S., Tarantal A.F, & Dandekar S. (2021). Gut germinal center regeneration and enhanced antiviral immunity by mesenchymal stem/stromal cells in SIV infection. JCI Insight, 6(12), e149033.

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Quantifying Virus-Specific T-Cell Responses

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Seven-week-old Ifnar1^−/− mice were infected with 1 × 10⁵ FFU of Z7 (G10) or PBS (control). Mice were euthanized on D8 p.i. to collect splenocytes. The splenocytes (3 × 10⁶) were plated in 6-well plates and re-stimulated with 0.1 MOI of Z1 for 24 h at 37 °C. During the final 8 h of the stimulation, Brefeldin A solution (BD Bioscience) was added in 1:1000 to block cytokine secretion. Cells were then stained with antibodies against mouse CD3 (FITC-conjugated, 0.25 µg/test, 1:200, BD Biosciences, catalog # 555274), CD4 (PerCp Cy 5.5-conjugated, 0.2 µg/test, 1:100, BD Biosciences, catalog # 550954), and CD8 (APC-conjugated, 0.2 µg/test, 1:100, BD Biosciences, catalog # 553035). Cells were fixed with 2% PFA overnight and permeabilized with 1 x Permeabilization buffer. Cells were then intracellularly stained with PE-Cy7 conjugated anti-IFNγ (0.2 µg/test, 1:100, BD Biosciences, catalog # 557649). Data were acquired using BD LSRFortessa^TM Cell Analyzer (BD Biosciences) and analyzed with FlowJo (v10.8.0.) software.

Nazneen F., Thompson E.A., Blackwell C., Bai J.S., Huang F, & Bai F. (2023). An effective live-attenuated Zika vaccine candidate with a modified 5′ untranslated region. NPJ Vaccines, 8, 50.

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Lymphocyte Isolation from Tissues

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Lamina propria, lung and liver lymphocytes were isolated as described previously (31 (link)). Briefly, lamina propria lymphocytes from intestine were isolated using the Lamina Propria Dissociation Kit (Miltenyi Biotec) according to the manufacturer’s instructions followed by Percoll gradient centrifugation. Lungs were digested with collagenase D and lymphocytes were isolated by Percoll gradient centrifugation. Liver lymphocytes were isolated by Percoll gradient centrifugation. Single cell suspensions were treated with Gey’s solution and resuspended in PBS added with 2% BSA. Antibodies used for flow cytometric analysis were as follows. Percp-Cy5.5-conjugated anti-CD4 and anti-CD62L, APC-Cy7-conjugated anti-CD8 and anti-CD44, PE-conjugated anti-H2Kb, anti-CD62L, RORγt and anti-Foxp3, PE-Cy7- conjugated anti-IFNγ, anti-CD69, anti-CD8, anti-CD25 and anti-CD44, Alexa-647- conjugated anti-TNFα, FITC-conjugated anti-CD44, anti-CD4 and anti-H2kb, APC-conjugated anti-phos-Erk, anti-H2Kd and anti-IL-17 antibodies were purchased from BD Biosciences or eBioscience. In some experiments, cells were stained with an Aqua dead cell exclusion dye. Foxp3 staining kit was from eBioscience. Samples were applied to LSRII flow cytometer (Becton Dickinson), and data were collected and analyzed using FACSDiva software (Becton Dickinson).

Luo L., Chen Y., Chen X., Zheng Y., Zhou V., Yu M., Burns R., Zhu W., Fu G., Felix J.C., Hartley C., Damnernsawad A., Zhang J., Wen R., Drobyski W.R., Gao C, & Wang D. (2020). Kras-deficient T cells attenuate graft-versus-host disease but retain graft-versus-leukemia activity. Journal of immunology (Baltimore, Md. : 1950), 205(12), 3480-3490.

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