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Il 4 pe cy7

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The IL-4 PE-Cy7 is a laboratory reagent used for the detection and analysis of interleukin-4 (IL-4) in biological samples. It is a fluorescently labeled antibody that binds to IL-4, allowing for its identification and quantification through flow cytometry or other immunoassay techniques.

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Lab products found in correlation

Cd25 apc, by BD (3 mentions) Perm wash, by BD (2 mentions) Il 10 pe, by BD (2 mentions) Lsr fortessa instrument, by BD (2 mentions) Fix perm buffer, by BD (2 mentions) Cd3 ecd, by Beckman Coulter (2 mentions) Ifnγ v450, by BD (2 mentions) Cd4 qdot655, by Thermo Fisher Scientific (2 mentions) Vα24 fitc, by Beckman Coulter (2 mentions) Withperm wash buffer, by Thermo Fisher Scientific (2 mentions) Pbs57 cd1d tet apc, by Beckman Coulter (2 mentions) Cd8 qdot 605, by Thermo Fisher Scientific (2 mentions) Tnf alexa700, by BD (2 mentions) Hla dr percp, by BD (2 mentions) Fix perm buffer, by Thermo Fisher Scientific (2 mentions) Cd38 pe, by BD (2 mentions) Il22 pe, by BD (2 mentions) Foxp3 pe, by BD (2 mentions) Ifn γ pe, by BD (2 mentions) Ly6g fitc, by BD (1 mentions)

Close spelling variants of this product

PE-Cy7 rat anti-mouse IL-4 (2 citations) Anti-IL-4 PE-Cy7 (2 citations) IL-4-PE (32 citations)

8 protocols using il 4 pe cy7

1

Multiparameter Flow Cytometry Analysis

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For surface staining, cells were stained with surface markers for 30
min on ice and washed twice with FACS buffer (PBS with 2% FBS and 2
mM EDTA buffer). After, cells were fixed and permeabilized with Fix/Perm
buffer (BD Biosciences) for 20 min on ice, washed twice with BD Perm/Wash,
and stained with the intracellular antibodies for 60 min on ice. For PoxP3
staining, cells were fixed and permeabilized with Fix/Perm Buffer
(eBiosciences, San Diego, CA, USA) for 60 min on ice, washed twice with
Perm/Wash buffer (eBiosciences) and stained with intracellular antibodies
for 60 min on ice. Subsequently, the cells were washed twice with the
respective Perm/Wash buffer and kept in 2% paraformaldehyde.
Antibodies used: PBS57-CD1d tet APC (kindly donated by NIH tetramer resource
facility), Vα24 FITC, and CD3 ECD were from Beckman Coulter
(Fullerton, CA), CD4 Qdot655, CD8 Qdot 605, and the viability marker AmCyan
were from Life Technologies (Carlsbad CA, USA), CD25 APC, CD38 PE, HLA-DR
PerCP, IFNγ V450, TNF Alexa700, IL-10 PE, and IL-4 PE-Cy7 were all
from BD bioscience. Data were acquired on a BD LSRFortessa instrument (BD
Biosciences) and analyzed using FlowJo Version 9.8.5 software (TreeStar,
Ashland, OR, USA).
Paquin-Proulx D., Ching C., Vujkovic-Cvijin I., Fadrosh D., Loh L., Huang Y., Somsouk M., Lynch S.V., Hunt P.W., Nixon D.F, & SenGupta D. (2016). Bacteroides are associated with GALT iNKT cell function and reduction of microbial translocation in HIV-1 infection. Mucosal immunology, 10(1), 69-78.
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2

Multiparametric Flow Cytometry Analysis

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LPLs were isolated and stimulated with Phorbol 12-myristate 13-acetate and ionomycin in the presence of brefeldin and monensin for 4 h at 37°C in a humidified CO2 incubator. Stimulated cells were stained for surface antigens with fluorescently labelled antibodies. Cells were then stained with fixable viability dye FV510 for live/dead discrimination followed by intracellular staining using Fix/Perm transcription factor kit (BD Biosciences, Franklin Lakes, NJ) and fluorescently labeled antibodies as follows: IL4 - PE-Cy7 (560699; BD Pharmingen, Franklin Lakes, NJ), KLRG1 – PerCP-Cy5.5 (563595; BD Pharmingen), IL22 – PE (BD Pharmingen; 516404), IFNγ- APC-Cy7 (561479; BD Pharmingen), Ly6G- FITC (BD Pharmingen; 561105), NK1.1 -FITC (561082; BD Pharmingen), CD3- FITC (561798; BD Pharmingen), CD19- FITC (BD Pharmingen; 561740), B220- FITC (561877; BD Pharmingen), NKp46- AF700 (561169; BD Pharmingen), RORγt- AF647 (562682; BD Pharmingen), viability dye FV510- BV510 (BD Pharmingen; 564406), CD127- BV421 (566377; BD Pharmingen), and CD45- BV786 (565477; BD Pharmingen). For Tregs quantification, Mouse Th17/Treg Phenotyping Kit (BD Biosciences) was used. Data were acquired on BD FACSCanto flow cytometer and analyzed using FlowJo software (FlowJo, Ashland OR).
Leon-Coria A., Kumar M., Workentine M., Moreau F., Surette M, & Chadee K. (2020). Muc2 Mucin and Nonmucin Microbiota Confer Distinct Innate Host Defense in Disease Susceptibility and Colonic Injury. Cellular and Molecular Gastroenterology and Hepatology, 11(1), 77-98.
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3

Flow Cytometry Immunophenotyping Protocol

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Cell phenotype was assessed by flow cytometry (BD LSRFortessa; BD Biosciences) after incubation with the following fluorescein-labeled antibodies: CD45-allophycocyanin (APC)-eFluor780, CD3e-fluorescein isothiocyanate (FITC), CD4-APC, CD8a-phycoerythrin (PE)-cy7, CD25-PE-cy7, Foxp3-PE, IFN-γ-AlexaFluor488, IL-4-PE-cy7, CD107a-PE, granzymeB-peridinin chlorophyll (PerCP)-eFluor710, perforin-FITC, CD11c-PE-cy7, CD80-PerCP-eFluor710, CD83-FITC, B7-H4-PE, and CD31-PE. All antibodies were purchased from eBioscience (San Diego, CA, USA), except CD4-APC, CD8a-PE-cy7, and Foxp3-PE (BD Biosciences). Intracellular antigens were determined after incubation with ionomycin (500 ng/mL; Abcam, Cambridge, UK) and phorbol-12-myristate13-acetate (10 ng/mL; Sigma-Aldrich, St. Louis, MO, USA) for 1 h and monensin (2 μM; eBioscience) for an additional 4 h. Fixation and permeabilization were performed prior to antibody incubation, and data were analyzed using FlowJo software (TreeStar, Ashland, OR, USA).
Yang M., Zhang Z., Chen J., Xu M., Huang J., Wang M., Li W., Wan X., Yuen M.F., Luo X., Xi D, & Ning Q. (2019). Soluble fibrinogen-like protein 2 promotes the growth of hepatocellular carcinoma via attenuating dendritic cell-mediated cytotoxic T cell activity. Journal of Experimental & Clinical Cancer Research : CR, 38, 351.
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4

Activation and Phenotyping of PBMCs

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PBMCs were cultured for 96h under different experimental conditions. Cells were counted, aliquoted and directly stained with 20μl (without fixation and permeabilization, as indicated by the manufacturer) with a CD8/CD69/CD3 antibody cocktail (BD FastImmune, BD Biosciences, cat# 340367) for analysis of activation. For immunophenotyping of the Th1/Th2/Th17 response, a different aliquot of cells was washed, fixed and permeabilized with Cytofix/Cytoperm (BD Biosciences) and stained with CD4-Percp-Cy5.5, 20μl test (cat# 341654), IFN-g-FITC, 5 μl/sample (cat# 554700), IL-4-PE-Cy7, 5 μl/sample (cat# 560672) and IL-17A- PE, 5 μl/sample (cat# 560486). Tregs were identified in a third aliquot of PBMCs that was washed, fixed and permeabilized with Human FOXP3 Buffer Set (BD Biosciences - cat# 560098), and stained with CD4-Percp-Cy5.5, 20 μl/sample (cat# 341654), FoxP3-AlexaFluor 488, 20 μl/sample (cat# 560047) and CD25-PE, 20 μl/sample (cat# 555432; all BD Biosciences). A minimum of 10,000 events were acquired in a FACSVerse flow cytometer and analyzed with FacSuite software (BD Biosciences).
de Medeiros M.C., Banerjee R., Liu M., Anovazzi G., D’Silva N.J, & Junior C.R. (2017). HNSCC subverts PBMCs to secrete soluble products that promote tumor cell proliferation. Oncotarget, 8(37), 60860-60874.
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5

Multiparametric Flow Cytometry Analysis of Murine Splenocytes

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The method of mouse euthanasia was described previously. Single cells from the spleen were suspended in RPMI 1640 culture medium (HyClone, Logan, UT, USA) supplemented with 10% fetal bovine serum (Gibco, Gaithersburg, MD, USA) and Cell Stimulation Cocktail (eBioscience, San Diego, CA, USA) and then cultured in 6-well plates for 5 hours in a 37°C/5% CO2 incubator.
Lymphocytes were separated from the spleen by passing through a 70 μm nylon mesh. Erythrocytes were lysed with ammonium chloride buffer, after which the remaining cells were washed and counted using a hemocytometer. One million cells were suspended in staining buffer with the following fluorochrome-conjugated antibodies: CD4-FITC (mAb; BD, Franklin Lakes, NJ, USA), CD25-APC (mAb; BD, Franklin Lakes, NJ, USA), IL-4-PECy7 (mAb; BD, Franklin Lakes, NJ, USA), IFN-γ-PE (mAb; BD, Franklin Lakes, NJ, USA), and Foxp3-PE (mAb; BD, Franklin Lakes, NJ, USA). Intracellular Foxp3, IL-4, and IFN-γ within lymphocytes were stained using a fixation/permeabilization staining kit (eBioscience, San Diego, CA, USA). All samples were analyzed using a FACSCanto II flow cytometer (BD, Franklin Lakes, NJ, USA) and analyzed using Flow Jo analysis software (Tree Star Inc.).
Liu Y., Yang M., Cui Y., Yao Y., Liao M., Yuan H., Gong G., Deng S, & Zhao G. (2020). A novel prevascularized tissue-engineered chamber as a site for allogeneic and xenogeneic islet transplantation to establish a bioartificial pancreas. PLoS ONE, 15(12), e0234670.
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6

Comprehensive Immunological Assays for In-Depth Analysis

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Gum arabic, activated carbon, 2,7-dichlorofluorescein diacetate (DCFDA), toluidine blue, protease inhibitor cocktail tablets, phosphatase inhibitor cocktail tablets, MILLIPLEX MAP Mouse Cytokine/Chemokine Magnetic Bead Panel were purchased from Merck KGaA (Darmstadt, Germany). 25% Aqueous Solution Glutaraldehyde, Paraformaldehyde, Sorensen's Phosphate Buffer, 2% Aqueous Solution Osmium Tetroxide, Ethyl Alcohol, Acetone, Araldite, Dibutyl phthalate (DBP) were purchased Electron Microscopy Sciences (Hatfield, USA). Leukocyte Activation Cocktail with BD GolgiPlug™, FACS antibodies include anti-mouse I-A/I-E-BV510, IgG1-BB700, IgM-BV605, IgE-BV786, IgD-BV711, CD1d-BV421, CD5-PE, CD45-BUV395, CD19-APC, CD45R/B220-BUV496, CD45-APC-Cy7, CD3e-FITC, CD4-V450, CD8-BV510, CD25-BV605, IL-4-PE-Cy7, IFN-γ-PE, FoxP3-AF647, CD103-BUV395, F4/80-BV711, CD80-BV650, CD11b-BV510, Ly6-G-PerCP Cy5.5, PE-Ly6-C, CD45-APC-Cy7, CD11c--AF700 were purchased from BD (Heidelberg, Germany); Another FACS antibody Anti-mouse-IL-17A-BV650 was purchased from eBioscience (Frankfurt am Main, Germany). Anti-mouse HMGB1, Anti-mouse phospho-p65, anti-mouse phospho-p38, anti-mouse GAPDH and anti-rabbit IgG HRP were obtained from Cell signaling Technology (Frankfurt am Main, Germany). IgE and histamine ELISA kits were purchased from Abcam (Berlin, Germany).
Lin S., Kühn F., Schiergens T.S., Zamyatnin AA J.r., Isayev O., Gasimov E., Werner J., Li Y, & Bazhin A.V. (2021). Experimental postoperative ileus: is Th2 immune response involved?. International Journal of Medical Sciences, 18(13), 3014-3025.
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7

Multiparameter Flow Cytometry Analysis

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For surface staining, cells were stained with surface markers for 30
min on ice and washed twice with FACS buffer (PBS with 2% FBS and 2
mM EDTA buffer). After, cells were fixed and permeabilized with Fix/Perm
buffer (BD Biosciences) for 20 min on ice, washed twice with BD Perm/Wash,
and stained with the intracellular antibodies for 60 min on ice. For PoxP3
staining, cells were fixed and permeabilized with Fix/Perm Buffer
(eBiosciences, San Diego, CA, USA) for 60 min on ice, washed twice with
Perm/Wash buffer (eBiosciences) and stained with intracellular antibodies
for 60 min on ice. Subsequently, the cells were washed twice with the
respective Perm/Wash buffer and kept in 2% paraformaldehyde.
Antibodies used: PBS57-CD1d tet APC (kindly donated by NIH tetramer resource
facility), Vα24 FITC, and CD3 ECD were from Beckman Coulter
(Fullerton, CA), CD4 Qdot655, CD8 Qdot 605, and the viability marker AmCyan
were from Life Technologies (Carlsbad CA, USA), CD25 APC, CD38 PE, HLA-DR
PerCP, IFNγ V450, TNF Alexa700, IL-10 PE, and IL-4 PE-Cy7 were all
from BD bioscience. Data were acquired on a BD LSRFortessa instrument (BD
Biosciences) and analyzed using FlowJo Version 9.8.5 software (TreeStar,
Ashland, OR, USA).
Paquin-Proulx D., Ching C., Vujkovic-Cvijin I., Fadrosh D., Loh L., Huang Y., Somsouk M., Lynch S.V., Hunt P.W., Nixon D.F, & SenGupta D. (2016). Bacteroides are associated with GALT iNKT cell function and reduction of microbial translocation in HIV-1 infection. Mucosal immunology, 10(1), 69-78.
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8

Characterization of T Helper Cells

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Spleen cells were treated with APC (10 µg/mL) for 24 h. 5 h prior to termination of the experiments, cells were stimulated with PMA (20 ng/mL)/Ionomycin (1 mM) in the presence of Monensin (5 µM). After stimulation, cells were collected and stained using a mouse antibody panel including CD3-AF700, CD4-BV711, IFN-γ-BV786, IL-4-PECY7, IL-17-BV421 and IL-22 (PE) (BD Biosciences, Franklin Lakes, NJ, USA) to detect Th cytokines in CD3+ and CD4+ lymphocytes. For specific Th cell phenotype identification, cells were treated with APC for 24 h and then stained with an antibody panel containing CD3-AF700, CD4-BV711, T-bet-BV786 (Th1), GATA3-PECY7 (Th2), Roγ-BV421 (Th17), FOX3-PE and CD25-APC (Treg) (BD Biosciences). Detection was performed using a BD LSR Fortessa flow cytometer (BD Biosciences).
Xue M., Lin H., Zhao R., Fryer C., March L, & Jackson C.J. (2022). Activated Protein C Protects against Murine Contact Dermatitis by Suppressing Protease-Activated Receptor 2. International Journal of Molecular Sciences, 23(1), 516.
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