Cd3 percp

Manufactured by BD

Sourced in United States, Denmark

The CD3-PerCP is a fluorochrome-conjugated antibody that binds to the CD3 cell surface antigen, which is expressed on T cells. It is a laboratory reagent used in flow cytometry applications for the identification and enumeration of T cells in biological samples.

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Lab products found in correlation

Cd8 pe, by BD (14 mentions) Cd56 apc, by BD (12 mentions) Cd4 fitc, by BD (12 mentions) Cd8 fitc, by BD (11 mentions) Cd25 pe cy7, by BD (8 mentions) Facscanto 2, by BD (8 mentions) Cd3 fitc, by BD (7 mentions) Ionomycin, by Merck Group (7 mentions) Cd8 apc h7, by BD (7 mentions) Trucount tube, by BD (6 mentions) Facscalibur, by BD (6 mentions) Cd14 fitc, by BD (5 mentions) Cd45ro apc, by BD (5 mentions) Cd45ra fitc, by BD (5 mentions) Cd4 apc h7, by BD (5 mentions) Cd4 pe cy7, by BD (5 mentions) Cd27 fitc, by BD (5 mentions) Ifn γ fitc, by BD (5 mentions) Il 17 apc, by R&D Systems (5 mentions) Cd19 pe cy7, by BD (5 mentions)

82 protocols using cd3 percp

Comprehensive Immune Cell Profiling

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The total number of WBCs, neutrophils, eosinophils, lymphocytes, and
monocytes were recorded by standard procedures using the ABX Pentra DX 120
(Horiba, United Kingdom/Germany). Furthermore, T-cell (CD3), T-helper cells
(CD4), T-cytotoxic cells (CD8), B-lymphocytes (CD19), NK (CD16/56) cells and
CD4+ recent thymic emigrants (CD4-RTE) absolute counts were performed by a
single-platform no-lyse-no-wash procedure. Fifty μl EDTA anti-coagulated
peripheral blood were incubated in TRUCount tubes (BD Biosciences, Denmark) with
a panel of conjugated monoclonal antibodies. The following combinations of
antibodies were used to characterize T cells as CD4 T cells (CD3-PerCP (clone
SK7), CD4-FITC (clone SK3), CD8 T cells (CD3-PerCP (clone SK7), CD8-PE (clone
SK1)) and CD4-RTE (recent thymic immigrant) cells as (CD3-ECD (clone UCHT1),
CD4-PC7 (clone SFCT12T4D11), CD31-PE (clone WM59), CD45RA-FITC (clone L48), and
CD45RO-PC5 (clone UCHL1), NK cells as (CD45-PerCP (clone 2D1), CD16/56-PE (clone
B73.1 + NCAM16.2), and CD3-FITC (clone SK7)), and B cells as (CD19-PE (clone
4G7), CD45-PerCP (clone 2D1)) (BD Biosciences, Beckman Coulter and AbD Serotec,
Denmark). The samples were measured on FC500 flow cytometer (Beckman Coulter,
Denmark). The laboratory participates in the quality assurance program by
National External Quality Assessment Site (NEQAS).

Oulhote Y., Shamim Z., Kielsen K., Weihe P., Grandjean P., Ryder L.P, & Heilmann C. (2016). Children’s white blood cell counts in relation to developmental exposures to methylmercury and persistent organic pollutants. Reproductive toxicology (Elmsford, N.Y.), 68, 207-214.

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Multicolor Flow Cytometry of Macaque PBMCs

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Macaque PBMCs (2 × 10⁶) isolated from peripheral blood were stained to determine T cell, B cell, and monocyte populations. PBMCs were washed with Roswell Park Memorial Institute Media (RPMI)-1640 + 10% FBS and resuspended in 100 μL of PBS. The cells were stained with antibodies (1 μg) for 30 min at room temperature with the following antibody panel (panel 1; Supplementary Figure S1): AmCyan-Live/Dead (Invitrogen, Grand Island, NY, USA); Qdot 655-CD8; PerCP-CD3 (BD Biosciences, San Jose, CA, USA); PE-CD45; Pacific Blue-CD14; APC-Cy7-CD4; PE-Cy7-CD16 (Biolegend, San Diego, CA, USA); ECD-CD20 (Beckman-Coulter, Brea, CA, USA) FITC-CD38 (StemCell Technologies, Vancouver, BC, Canada); APC-CD66 (Miltenyi Biotech, Auburn, CA, USA). PBMCs were stained for 30 min at room temperature to assess SIV co-receptor expression with the following panel (panel 2; Supplementary Figure S2): AmCyan-Live/Dead (Invitrogen, Grand Island, NY, USA); Qdot 655-CD8; PerCP-CD3 (BD Biosciences, San Jose, CA, USA); APC-Cy7-CD4 (Biolegend, San Diego, CA, USA); AF488-CXCR4 (R&D Systems); APC-CCR5 (BD Biosciences). Cells were washed with 2 mL of PBS, re-suspended in 300 μL of PBS + 1% paraformaldehyde (PFA), and acquired on an LSRII (BD Biosciences). Data were analyzed using FACS DIVA and FlowJo (TreeStar, Ashland, OR, USA) software.

McTernan P.M., Siggins R.W., Catinis A., Amedee A.M., Simon L, & Molina P.E. (2022). Chronic Binge Alcohol and Ovarian Hormone Loss Dysregulate Circulating Immune Cell SIV Co-Receptor Expression and Mitochondrial Homeostasis in SIV-Infected Rhesus Macaques. Biomolecules, 12(7), 946.

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Characterizing Liver NK Cell Activation and Degranulation

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Flow cytometry was performed using a BD LSRII flow cytometer and data were analysed on FACSDiva software (BD Biosciences). All antibodies comprised fluorescent-conjugated mouse monoclonal IgG1.
Liver NK cell CD69: washed LMCs were labelled with Fluorescein isothiocyanate (FITC)-CD56, PerCP-CD3 and PE-CD69 IgG1 antibodies (all BD Biosciences). Controls were labelled with PE-conjugated IgG1 isotype.
CD107 a/b degranulation: LMCs were incubated for 24 hours with 1 PFU/cell reovirus or PBS, washed twice in Hanks' Balanced Salt Solution (HBSS) (Sigma) and coincubated with target cells at a ratio of 5:1 for 4 hours. At 1 hour, cells were labelled with FITC-CD107a, FITC-CD107b antibodies (BD Biosciences), PE-CD56 (AbD Serotec) and PerCP-CD3 (BD Biosciences), with Brefeldin A solution (Biolegend).

Samson A., Bentham M.J., Scott K., Nuovo G., Bloy A., Appleton E., Adair R.A., Dave R., Peckham-Cooper A., Toogood G., Nagamori S., Coffey M., Vile R., Harrington K., Selby P., Errington-Mais F., Melcher A, & Griffin S. (2016). Oncolytic reovirus as a combined antiviral and anti-tumour agent for the treatment of liver cancer. Gut, 67(3), 562-573.

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Multiparametric Flow Cytometry of T-Lymphocytes

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T-lymphocytes were analyzed in PBMCs by nine-color flow-cytometry. PBMCs were incubated with the next surface-labeled monoclonal-antibodies, CD3-PercP, CCR7-PECY7 (Becton-Dickinson, San José, CA, USA), CD8-Alexa405, CD45RA-APC (Caltag, San Francisco, CA, USA) and CD27-APCAlexa780 (eBioscience, San Diego, CA, USA).
For intracytoplasmic staining, PBMCs were fixed and permeabilized (Fix and Perm, Caltag, San Francisco CA, USA), and cytokines were stained with IL-4-PE, IFNγ Alexa700 and IL-17A-FITC (Becton-Dickinson, San José, CA, USA). All samples were stained with a dead cell-discriminator simultaneously with antibody addition (fixable aqua dead cell stain kit for 405 nm excitation; Molecular Probes, Eugene, OR). Samples were acquired in a FacsAria-II flow-cytometer and were analyzed using FacsDiva 5.0 and Flow-Jo 10.0 software.

Alvarez-Mon M.A., Gómez-Lahoz A.M., Orozco A., Lahera G., Diaz D., Ortega M.A., Albillos A., Quintero J., Aubá E., Monserrat J, & Alvarez-Mon M. (2021). Expansion of CD4 T Lymphocytes Expressing Interleukin 17 and Tumor Necrosis Factor in Patients with Major Depressive Disorder. Journal of Personalized Medicine, 11(3), 220.

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Immunophenotypic Analysis of NK and T Cells

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Immunophenotypic analysis was performed using three colour flow cytometry on a FACS Calibur (Becton Dickinson) and six colour flow cytometry on a FACS Canto (Becton Dickinson). Antibodies used were CD45 FITC, 2DL2/3 FITC, CD56 PE, NKG2D PE, CD3 PerCP, Streptavidin PerCP, CD56 PE-Cy7, 2DL1 APC, CD56 APC, CD3 APC-Cy7, 3DL1 biotin (all Becton Dickinson). Ig isotype controls were used where appropriate. Lymphocyte gating was performed using the forward scatter versus side scatter plot and 10 000 events were acquired. NK cells were identified as CD3⁻CD56⁺, T cells as CD3⁺CD56⁻, and CD56⁺ T cells as CD3⁺CD56⁺. Data was analysed using Cell Quest/FACS Diva software.

Pugh S.A., Harrison R.J., Primrose J.N, & Khakoo S.I. (2014). T cells but not NK cells are associated with a favourable outcome for resected colorectal liver metastases. BMC Cancer, 14, 180.

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Flow Cytometric Characterization of T-Cell Subsets

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Similar to previous literature reports [23 (link)], the flow cytometry antibodies used in the following tests were purchased from BD Biosciences. BMMC was incubated with phorbol-12-myristate 13-acetate (PMA) and Ionomycin (Sigma, USA) for 4 h at 37 °C. The cells were incubated with flow cytometry antibodies against CD3-PerCP and CD8-APC (Becton Dickinson) for 15 min in dark. Then, the cells were treated with IntraPreP permeabilization Reagent B (Becton Dickinson) and were stained anti-human IFNγ-FITC and IL-4-PE (Becton Dickinson) for 15 min. After the cells were washed with cold PBS, an appropriate amount of 50ul PBS was added, flow cytometry was performed in FACS Calibur, and the results were analyzed by CellQuest software. Cell subsets are defined as follows: Th1 (CD8-INF-γ+), Th2 (CD8- IL-4+), Tc1 (CD8+ INF-γ+), and Tc2 (CD8+ IL-4+).

Zhang Z., Huang N., Xv F., Zhao S., Guo J., Zhao Y, & Chang C. (2022). Decreased FOXO1 Expression Is Correlated with Poor Prognosis in Myelodysplastic Syndromes. Current Oncology, 29(10), 6933-6946.

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Multicolor Flow Cytometry Analysis of NK Cells

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For measurement of phosphorylated pSTAT-4 20 min and for measurement of IFN-γ and perforin 18 h in vitro stimulated PBMC (1x10⁶/ml) from HC and MM patients were analyzed. Three hours before IFN-γ analysis, brefeldin A was added at concentration of 10 μg/ml. The cells were then stained with mAbs against cell surface molecules: CD3PerCP, CD56PE (for IFN-γ analysis) and CD56FITC (for pSTAT-4 and perforin analyses) (Becton Dickinson), followed by fixing and permeabilizing with the Permeabilizing Solution 2 (Perm 2) (Becton Dickinson) accords to the manufacturer’s instructions. Then the cells were incubated with anti-IFN-γFITC (clone 25723.11), anti-pSTAT-4PE (clone 38/p-Stata4) (Becton Dickinson), anti-Perforin RPE (clone deltaG9) (Invitrogen, Madison, USA) or IgG1 isotype control (Becton Dickinson). Finally, the cells were washed and analyzed by flow cytometry directly after preparation. The percentages and MFI of intracellular IFN-γ, pSTAT-4 and perforin were analyzed in CD3⁻CD56⁺ NK cells and their CD3⁻CD56^dim+ and CD3⁻CD56^bright+ subsets.

Mirjačić Martinović K., Babović N., Džodić R., Jurišić V., Matković S, & Konjević G. (2015). Favorable in vitro effects of combined IL-12 and IL-18 treatment on NK cell cytotoxicity and CD25 receptor expression in metastatic melanoma patients. Journal of Translational Medicine, 13, 120.

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Immune Cell Profiling After Transplantation

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T, B, and NK cell counts were determined in months + 1, + 2, + 3, and + 6 post-transplantation. Lymphocyte subsets in peripheral blood were quantified using Trucount Tubes (Becton Dickinson, Albertslund, Denmark) together with the following panel of conjugated monoclonal antibodies and analyzed on a FC500 flow cytometer (Beckman Coulter, Copenhagen, Denmark): CD3-PerCP, CD3-FITC, CD4-FITC, CD8-PE, CD45-PerCP, CD16/56-PE, CD20-FITC, and CD19-PE (Becton Dickinson). CD3⁺ T cells, CD3⁺CD4⁺ T cells, and CD3⁺CD8⁺ T cells were determined. NK cells were differentiated by CD3⁻CD45⁺CD16⁺CD56⁺ phenotype. The following B cell phenotypes were distinguished: total B cells (CD45⁺CD19⁺), mature B cells (CD45⁺CD19⁺CD20⁺), and immature B cells (CD45⁺CD19⁺CD20⁻). Data of these immune cell populations have been published previously in a different context [46 (link)].

Ingham A.C., Kielsen K., Cilieborg M.S., Lund O., Holmes S., Aarestrup F.M., Müller K.G, & Pamp S.J. (2019). Specific gut microbiome members are associated with distinct immune markers in pediatric allogeneic hematopoietic stem cell transplantation. Microbiome, 7, 131.

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Comprehensive T-Cell Immunophenotyping by Flow

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T-lymphocytes were studied in PBMCs by nine-color flow cytometry. PBMCs were incubated with the next surface-labeled monoclonal-antibodies, CD3-PercP, CCR7-PECY7 (Becton-Dickinson, BD, CA, USA), CD8-Alexa405, CD45RA-APC (Caltag, Carlsbad, CA, USA) and CD27-APCAlexa780 (eBioscience, San Diego, CA, USA).
For intracytoplasmic staining, cells were fixed and permeabilized (Fix and Perm, Caltag, Carlsbad, CA, USA), and cytokines were stained with IL-4-PE, IFNγ-Alexa700 and IL-17A-FITC (Becton-Dickinson, BD, CA, USA). All samples were stained with a dead cell-discriminator simultaneously with antibody addition (Fixable aqua dead cell stain kit for 405 nm excitation; Molecular Probes, Eugene, OR, USA).
Samples were acquired in a FacsAria-II flow cytometer and were analyzed using FacsDiva 5.0 and Flow-Jo 7.0 software (Becton-Dickinson, BD, San Jose, CA, USA).

Monserrat Sanz J., Bohórquez C., Gómez A.M., Movasat A., Pérez A., Ruíz L., Diaz D., Sánchez A.I., Albarrán F., Sanz I, & Álvarez-Mon M. (2020). Methrotexate Treatment Inmunomodulates Abnormal Cytokine Expression by T CD4 Lymphocytes Present in DMARD-Naïve Rheumatoid Arthritis Patients. International Journal of Molecular Sciences, 21(18), 6847.

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Activation of PBMCs and IL-17 Secretion

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PBMCs were activated for 24 hours with Dynabeads Human T-Activator CD3/CD28 (Life Technologies) and cultured for 3 hours with or without 20 µg/mL CD30/Fc chimera (R&D Systems) in RPMI + 10% FCS. Cells were then harvested and tested for IL-17 secretion using Cytokine Secretion Assays (Miltenyi Biotec, Bergisch Gladbach, Germany) following manufacturer's instructions. Briefly, cells were suspended in 2 mL medium, washed with 2 mL cold buffer at 300 ×g for 10 minutes, and resuspended in 90 μL cold medium. Cells were incubated 5 minutes on ice with 10 μL IL-17 catch reagent. Cells were diluted with warm medium and incubated for 45 minutes at 37°C, washed with cold buffer and then stained with CD3 PerCp (Becton Dickinson), CD30L PE (R&D Systems), and IL-17 Detection Antibody APC. Cells were acquired on a FACSCanto cytometer (Becton Dickinson). Analysis was performed with FlowJo 9.3.3 software (Tree Star).

Barbieri A., Dolcino M., Tinazzi E., Rigo A., Argentino G., Patuzzo G., Ottria A., Beri R., Puccetti A, & Lunardi C. (2015). Characterization of CD30/CD30L+ Cells in Peripheral Blood and Synovial Fluid of Patients with Rheumatoid Arthritis. Journal of Immunology Research, 2015, 729654.

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