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Cd4 af700

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The CD4-AF700 is a fluorochrome-conjugated antibody used for the detection and quantification of CD4+ T cells in flow cytometry analysis. It is designed to bind specifically to the CD4 surface antigen expressed on T helper cells. The fluorochrome attached to the antibody is Alexa Fluor 700, which can be excited by a red laser and detected in the near-infrared range.

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Lab products found in correlation

Cd39 pe cy7, by Thermo Fisher Scientific (2 mentions) Lsrii flow cytometer, by BD (2 mentions) Il 17 pe, by BD (1 mentions) Mhcii pacificblue, by BioLegend (1 mentions) Cd8 pacific blue, by Thermo Fisher Scientific (1 mentions) Cd25 apc, by Thermo Fisher Scientific (1 mentions) Cd45r b220 percp cy5, by BD (1 mentions) Ifn γ percp cy5, by BD (1 mentions) Foxp3 af488, by BD (1 mentions) Cd86 apc, by Thermo Fisher Scientific (1 mentions) Cd49b pe cy7, by Thermo Fisher Scientific (1 mentions) Cd11b pe cy7, by BD (1 mentions) F4 80 apc, by BioLegend (1 mentions) Cytofix cytoperm, by BD (1 mentions) C kit pe cy7, by Thermo Fisher Scientific (1 mentions) Lsrii sorp, by BD (1 mentions) Cd45 percp cy5, by BD (1 mentions) Anti il 4 fitc, by BD (1 mentions) Cd11b apc efluor780, by Thermo Fisher Scientific (1 mentions) Fortessa, by BD (1 mentions)

6 protocols using cd4 af700

1

Comprehensive Immune Cell Profiling by Flow Cytometry

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Flow cytometry was performed as previously described [38 (link)]. Briefly, cells were incubated with LIVE/DEAD Fixable cell stain (Invitrogen), then stained for 20 min with: MHCII-PacificBlue, F4/80-APC (Biolegend), CD4-AF700, CD8-PacificBlue, CD25-APC, CD86-APC, CD49b-Pe-Cy7 (eBioscience), CD11b-PE-Cy7, CD45R/B220-PerCP-Cy5.5 (BD-Pharmingen). For intracellular staining, the samples were stimulated for 3 h (100 ng/ml phorbol 12-myristate 13-acetate, 1 μg/ml ionomycin; monensin added after 1 h). Cells were fixed and permeabilized with Cytofix/Cytoperm (BD), then stained with IL-10-FITC, IFN-γ-PerCP-Cy5.5, IL-17-PE, FoxP3-AF488 (BD-Pharmingen) antibodies before analysis on LSRII flow cytometer (BD), collecting 100,000 live events. Data was analyzed using FlowJo (v7.6.5, Tree Star, Ashland, OR, USA).
Schuijs M.J., Hartmann S., Selkirk M.E., Roberts L.B., Openshaw P.J, & Schnoeller C. (2016). The Helminth-Derived Immunomodulator AvCystatin Reduces Virus Enhanced Inflammation by Induction of Regulatory IL-10+ T Cells. PLoS ONE, 11(8), e0161885.
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2

Quantitative Analysis of Immune Cells

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Stained cells were analyzed using a BD LSR II-SORP or Fortessa system (Becton Dickinson) and analyzed with FlowJo software (Tree Star). The following anti-mouse mAbs were used: CD45- PerCPCy5, CD45-APC, CD8-APC, anti-IFNγ-PECy7, and anti-IL-4-FITC (BD Biosciences), CD4-AF700, CD11b APC-eFluor780, and c-kit PECy7 all from (eBiosciences). Anti human CD2 (BioLegend) was used to detect IL-4 production in KN2 mice. To detect IL-4Rα on primary keratinocytes, the following anti-mouse mAbs were used: CD124, anti IL-4Rα-PE (BD, Pharmingen), and CD49f-Brillant Violet 421 (BioLegend). Imaging flow cytometry was performed using an ImageStream cytometer (Amnis, Millipore) at low speed. Data were analyzed with the IDEAS software.
Descatoire M., Hurrell B.P., Govender M., Passelli K., Martinez-Salazar B., Hurdayal R., Brombacher F., Guler R, & Tacchini-Cottier F. (2017). IL-4Rα Signaling in Keratinocytes and Early IL-4 Production Are Dispensable for Generating a Curative T Helper 1 Response in Leishmania major-Infected C57BL/6 Mice. Frontiers in Immunology, 8, 1265.
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3

Multiparametric Flow Cytometry Analysis

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BALF cells and PBMCs were stained for extra- and intracellular markers using the following antibodies: CD3-APC-eFluor780(SK7), CD4-AF700(OKT4), CD45RO-FITC(UCHl-1), CD95-APC(DX2), FoxP3-APC(PCH101) (eBiosciences) and CD25-PE(M-A251), CD25-PE-Cy7(M-A251), CD127-V450(hIL7R-M21) anti-CTLA-4-BV421(BNI3) (BD biosciences) and CD45RA-PE-Texas Red(MEM-56) (Invitrogen). Fixable Aqua Dead Cell Stain kit for 405 nm (Invitrogen, Molecular Probes) was used as live-dead marker. Cells were measured on a Flow cytometer LSRII (BD Biosciences).
Broos C.E., van Nimwegen M., Kleinjan A., ten Berge B., Muskens F., in ’t Veen J.C., Annema J.T., Lambrecht B.N., Hoogsteden H.C., Hendriks R.W., Kool M, & van den Blink B. (2015). Impaired survival of regulatory T cells in pulmonary sarcoidosis. Respiratory Research, 16(1), 108.
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4

Identification of CD8+ T cells in Tumor-Bearing Kidney Tissue

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To identify the CD8+ T cells among the CD45.2+ leukocytes located in the tumor-bearing kidney tissue, intravascular staining was done using anti-CD45.2-PE (BioLegend, San Diego, CA) (18 (link)). Tumor-bearing kidneys were harvested, manually disrupted, and digested with DNaseI (15 µg/ml; Sigma) and Liberase Blendzyme 3 (0.026 Wuensch unites/ml; Roche). After blocking with anti-CD16/32 in normal mouse serum, cells were stained (anti-CD45.2-BV650, CD3-v500, CD4-AF700, CD8-BUV395; eBioscience, San Diego, CA or BioLegend) and analyzed using multi-parameter flow cytometry on a BD LSR Fortessa (BD Biosciences, San Diego, CA) and FlowJo software (TreeStar Inc., Ashland, OR).
Murphy K.A., James B.R., Sjaastad F.V., Kucaba T.A., Kim H., Brincks E.L., Chua SC J.r., Wilber A, & Griffith T.S. (2018). Elevated leptin during diet-induced obesity reduces the efficacy of tumor immunotherapy. Journal of immunology (Baltimore, Md. : 1950), 201(7), 1837-1841.
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5

Comprehensive Immunophenotyping of PBL

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The following anti-human monoclonal antibodies (mAb) were used for flow cytometry staining: CD8-FITC, CD137-PE, CD39-PE-Cy7, CD4-AF700, LAG3-PE, PD-1-PE, PD-L1-PE (all eBioscience. Waltham, USA), BLTA-BV421, GITR-BV421, OX40-PE-Cy5, TIM3-PB (all Biolegend, London, UK), and CD45-AMCyan (BD Bioscience, San Jose, CA, USA). All antibodies were pre-titrated using stimulated and non-stimulated PBL to determine optimal staining dilutions. For IHC, PD-L1 (E1L3N) XP Rabbit mAb was purchased from Cell Signaling Technology, Danvers, MA, USA.
Puntigam L.K., Jeske S.S., Götz M., Greiner J., Laban S., Theodoraki M.N., Doescher J., Weissinger S.E., Brunner C., Hoffmann T.K, & Schuler P.J. (2020). Immune Checkpoint Expression on Immune Cells of HNSCC Patients and Modulation by Chemo- and Immunotherapy. International Journal of Molecular Sciences, 21(15), 5181.
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6

Multiparametric Flow Cytometry of Immune Cells

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The following anti-human monoclonal antibodies (mAb) were used for flow cytometry: CD4 AF700 (Cat 56-0048-82), CD73 FITC (Cat 11-0739-42), CD73 PE (Cat 12-0739-42), CD39 PE-Cy7 (Cat 25-0399-42), PD1 (CD279) PE (Cat 12-2799-42), and CCR7 (CD197) PE-Cy7 (Cat 25-1979-42) mAb (all eBioscience, San Diego, CA); CD5 AF700 (Cat 561159), CD19 PE-Cy7 (Cat 555414), and IgM PE (Cat 555783) (all BD Pharmingen, Heidelberg, Germany); CD26 APC (Cat 302710), CD25 FITC (Cat 356105), and CD40 PE-Cy7 (Cat 334322) (all Biolegend, San Diego, CA); CD25 PE (Cat 130-101-426) and IgA APC (Cat 130-093-113) (Milenty Biotec, Bergisch Gladbach, Germany). All mAb were titrated using normal PBMC to establish optimal staining dilutions.
Ziebart A., Huber U., Jeske S., Laban S., Doescher J., Hoffmann T.K., Brunner C., Jackson E.K, & Schuler P.J. (2017). The influence of chemotherapy on adenosine-producing B cells in patients with head and neck squamous cell carcinoma. Oncotarget, 9(5), 5834-5847.
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