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Percp cy5.5 conjugated anti cd4

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PerCP-Cy5.5-conjugated anti-CD4 is a fluorescently labeled antibody that binds to the CD4 cell surface receptor. It is a tool used for the detection and analysis of CD4+ cells in flow cytometry applications.

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Lab products found in correlation

Live dead fixable aqua, by Thermo Fisher Scientific (3 mentions) Lsrfortessa x 20 flow cytometry, by BD (3 mentions) Fitc conjugated anti pd 1, by BioLegend (3 mentions) Apc cy7 conjugated anti cd3, by BD (3 mentions) Facscalibur, by BD (3 mentions) Pe cy7 conjugated anti cd45ra, by BD (2 mentions) Brilliant violet 421 conjugated anti cd25, by BD (2 mentions) Brilliant violet 605 conjugated anti cd8, by BioLegend (2 mentions) Apc conjugated anti cd127, by BioLegend (2 mentions) Pe conjugated foxp3 antibody, by Thermo Fisher Scientific (2 mentions) Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (2 mentions) Pe conjugated anti cd3, by BioLegend (2 mentions) Fitc conjugated anti cd25, by BioLegend (1 mentions) Fitc conjugated anti ifn γ, by BioLegend (1 mentions) Percp cy5.5 conjugated anti cd8, by BioLegend (1 mentions) Apc conjugated anti il 17, by BioLegend (1 mentions) Facscalibur flow cytometer, by BD (1 mentions) Isotype matched control antibody, by BioLegend (1 mentions) Fitc conjugated anti f4 80, by BioLegend (1 mentions) Anti cd29 hmβ1 1, by BioLegend (1 mentions)

7 protocols using percp cy5.5 conjugated anti cd4

1

Multi-color Flow Cytometry of PBMCs

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Multi-color flow cytometry of PBMCs was performed at CNUH. The following human antibodies were used for multi-color flow cytometry: Brilliant Violet 421-conjugated anti-CD25, PE-Cy7 conjugated anti-CD45RA, and APC-Cy7 conjugated anti-CD3 (all from BD Biosciences, San Jose, CA, USA) and Brilliant Violet 605-conjugated anti-CD8, PerCP-Cy5.5-conjugated anti-CD4, FITC-conjugated anti-PD-1, and APC-conjugated anti-CD127 (all from BioLegend, San Diego, CA, USA). For intracellular staining of FOXP3, cells were fixed and permeabilized after surface staining using the Foxp3/Transcription factor staining buffer set (eBioscience) and incubated with PE-conjugated FOXP3 antibody (eBioscience, San Diego, CA, USA). To exclude dead cells, single-cell suspensions were first incubated for 20 min in viability dye (LIVE/DEAD Fixable Aqua, Thermo Fisher). Stained cells were analyzed using BD LSR Fortessa X-20 flow cytometry (BD Biosciences). Fluorescence-activated cell sorting (FACS) analysis was performed using FlowJo software (Tree Star, Ashland, OR, USA).
Kang D.H., Chung C., Sun P., Lee D.H., Lee S.I., Park D., Koh J.S., Kim Y., Yi H.S, & Lee J.E. (2021). Circulating regulatory T cells predict efficacy and atypical responses in lung cancer patients treated with PD-1/PD-L1 inhibitors. Cancer Immunology, Immunotherapy, 71(3), 579-588.
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2

Profiling T Cell Phenotypes and Cytokines

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We isolated mononuclear cells from fresh LN samples, as previously described [28 (link)]. Phenotypic & cytokine analysis of T cells was performed using FACSCalibur flow cytometer (BD biosciences, USA). The following mouse anti-human monoclonal antibodies (mAbs) were used: PerCp-Cy5.5 conjugated anti-CD4 (Clone: RPA-T4), PerCp-Cy5.5 conjugated anti-CD8 (Clone: SK1), Fluorescein isothiocyanate (FITC)-conjugated anti-CD25 (Clone: M-A251), Allophycocyanin (APC)-conjugated anti-CD127 (Clone: A019D5), FITC-conjugated anti-IFN-γ (Clone: 4S.B3), PE anti-human TGF-β (Clone: TW4-9E7), APC-conjugated anti-IL-17 (Clone: BL168), APC-conjugated anti-IL-4 (Clone: 8D4-8), APC-conjugated anti-IL-10 (Clone: JES3-19F1), and their respective isotype controls were purchased from Biolegend, USA. Phycoerythrin (PE)-conjugated anti-Foxp3 (Clone: 259 D/C7), PE-conjugated anti-tumor necrosis factor-α (TNF-α) (Clone: MAB11) and their isotype controls were obtained from BD biosciences.
Norouzian M., Mehdipour F., Ashraf M.J., Khademi B, & Ghaderi A. (2022). Regulatory and effector T cell subsets in tumor-draining lymph nodes of patients with squamous cell carcinoma of head and neck. BMC Immunology, 23, 56.
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3

Characterization of Immune Cells and Cytokine Levels

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Flow cytometry was performed using FACS Calibur (BD Biosciences). CT26 cells were cultured with or without 50 ng/mL interferon (IFN)-γ (Tonbo Biosciences) for 48 hours and stained with anti-CCR7 (clone 4B12), anti-PD-L1 (clone 10F.9G2), and isotype-matched control antibodies (BioLegend). iFib, iMSC, and iMSC/CCL19 were stained with the following antibodies as described previously (3): anti-PDGFRα (clone APA5), anti-PDGFRβ (clone APB5), anti-CD34 (clone HM34), anti-Sca-1 (clone D7), and anti-CD29 (HMβ1–1) (all antibodies from BioLegend). Anti-CD44 (clone IM7), anti-CD45 (clone 30-F11), and anti-CD117 (clone ACK2) antibodies were purchased from Tonbo Biosciences. Cell suspensions from tumor or spleen were stained with the following antibodies: PE-conjugated anti-CD3, PerCP-Cy5.5-conjugated anti-CD4, FITC-conjugated anti-CD8α, PerCP-Cy5.5-conjugated anti-CD11c, APC-conjugated anti-CCR7, FITC-conjugated anti-PD-1, and FITC-conjugated anti-F4/80 (antibodies purchased from BioLegend).
ELISA iFib, iFib/CCL19, iMSC, and iMSC/CCL19 cells were cultured for 48 hours. Then, the supernatants were collected, and the levels of CCL19 in the supernatants were measured using the mouse MIP3 beta ELISA Kit (#ab100729, Abcam) and Multiskan FC Basic plate-reader at 450 nm (Thermo Fisher).
Iida Y., Yoshikawa R., Murata A., Kotani H., Kazuki Y., Oshimura M., Matsuzaki Y, & Harada M. (2020). Local injection of CCL19-expressing mesenchymal stem cells augments the therapeutic efficacy of anti-PD-L1 antibody by promoting infiltration of immune cells. Journal for Immunotherapy of Cancer, 8(2), e000582.
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4

Multicolor Flow Cytometry Analysis

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Cells were resuspended in PBS containing 2% FCS (v/v), pretreated with anti-CD16/32 (93; BioLegend) and then stained with optimal amounts of FITC-conjugated anti-CD45.1, PerCP-Cy5.5-conjugated anti-CD4 and PE-conjugated anti-CD3 (all Biolegend). Cells were analyzed on FACSCalibur (BD Biosciences) or Attune NxT Flow Cytometer (Thermo Fisher Scientific). Data analysis was performed using FlowJo software (Tree Star).
Frick R., Høydahl L.S., Petersen J., du Pré M.F., Kumari S., Berntsen G., Dewan A.E., Jeliazkov J.R., Gunnarsen K.S., Frigstad T., Vik E.S., Llerena C., Lundin K.E., Yaqub S., Jahnsen J., Gray J.J., Rossjohn J., Sollid L.M., Sandlie I, & Løset G.Å. (2021). A high-affinity human TCR-like antibody detects celiac disease gluten peptide-MHC complexes and inhibits T-cell activation. Science immunology, 6(62), eabg4925.
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5

Multi-color Flow Cytometry for PBMC Profiling

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Multi-color flow cytometry of PBMCs was performed at CNUH. The following human antibodies were used for multi-color flow cytometry: Brilliant Violet 421-conjugated anti-CD25, PE-Cy7 conjugated anti-CD45RA, and APC-Cy7 conjugated anti-CD3 (BD Biosciences, San Jose, CA, USA), and Brilliant Violet 605-conjugated anti-CD8, PerCP-Cy5.5-conjugated anti-CD4, FITC-conjugated anti-PD-1, and APC-conjugated anti-CD127 (BioLegend, San Diego, CA, USA). For intracellular staining of FOXP3, cells were fixed and permeabilized after surface staining using the Foxp3/Transcription factor staining buffer set (eBioscience) and incubated with a PE-conjugated FOXP3 antibody (eBioscience, San Diego, CA, USA). To exclude dead cells, single-cell suspensions were first incubated for 20 min in a viability dye (LIVE/DEAD Fixable Aqua, Thermo Fisher). The stained cells were analyzed using BD LSR Fortessa X-20 flow cytometry (BD Biosciences). Fluorescence-activated cell sorting was performed using FlowJo software (Tree Star, Ashland, OR, USA).
Hong G., Sun P., Chung C., Park D., Lee S.I., Kim N., Lee S.E., Lee J.E., Kang Y.E, & Kang D.H. (2022). Plasma GDF15 levels associated with circulating immune cells predict the efficacy of PD-1/PD-L1 inhibitor treatment and prognosis in patients with advanced non-small cell lung cancer. Journal of Cancer Research and Clinical Oncology, 149(1), 159-171.
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6

Multicolor Flow Cytometry of PBMCs

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Multicolor flow cytometry of PBMCs was performed at CNUH. The following human antibodies were used for multi‐color flow cytometry: APC‐Cy7‐conjugated anti‐CD3, PE‐CF594‐conjugated anti‐PD1 (both from BD Pharmingen), PE‐conjugated anti‐CXCR3 (CD183), PerCP‐Cy5.5‐conjugated anti‐CD4, APC‐conjugated anti‐CCR6 (CD196), BV421‐conjugated anti‐CCR4 (CD194), and BV605‐conjugated anti‐CD8 (all from BioLegend). To exclude dead cells, single‐cell suspensions were first incubated for 20 min in viability dye (LIVE/DEAD Fixable Aqua, Thermo Fisher Scientific). Stained cells were analyzed using BD LSR Fortessa X‐20 flow cytometry (BD Biosciences). Fluorescence‐activated cell sorting (FACS) analysis was performed using FlowJo software (Tree Star).
Kang D.H., Choi S., Sun P., Chung C., Park D., Lee S., Koh J.S., Kim Y, & Lee J.E. (2022). The rest period between chemotherapy and immunotherapy influences the efficacy of immune checkpoint inhibitors in lung cancer. Thoracic Cancer, 13(16), 2346-2354.
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7

Multicolor Flow Cytometry Analysis

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Cell suspensions from spleens were prepared by grinding tissue through sterile wire mesh and stained with a panel of uorescent-conjugated antibodies including PerCP/Cy5.5 conjugated anti-B220 (Biolegend, 103236), FITC-conjugated anti-FAS (BD Pharmingen, 554257), eFluor 660-conjugated anti-GL7 (Invitrogen, 50-5902-82), PerCP/Cy5.5 conjugated anti-CD4 (Biolegend, 100434 ), PE-conjugated anti-CXCR5 (Biolegend, 145504), and APC-conjugated anti-PD1 (Biolegend, 109112). Samples were run on a FACS Calibur (BD), and the data were analyzed with FlowJo software (TreeStar, Portland, OR).
Xing Y., Guo W., Wu M., Xie J., Huang D., Hu P., Zhou M., Zhang L., Zhang Q., Wang P., Wang X., Wang G., Wu H., Zhou C., Chen Y., Liu M., Yi Z, & Sun Z. (2022). An orally available small molecule BCL6 inhibitor effectively suppresses diffuse large B cell lymphoma cells growth in vitro and in vivo. Cancer letters, 529.
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