Cd3 bv650

Manufactured by BD

Sourced in China, United States

CD3-BV650 is a fluorochrome-conjugated antibody that binds to the CD3 antigen, which is expressed on the surface of T cells. It is designed for use in flow cytometry applications to identify and characterize T cell populations.

Automatically generated - may contain errors

Lab products found in correlation

Cd27 pe cy7, by BioLegend (2 mentions) Facsdiva software, by BD (2 mentions) Brefeldin a, by BD (2 mentions) Live dead aqua, by Thermo Fisher Scientific (2 mentions) Live dead fixable near ir stain, by Thermo Fisher Scientific (1 mentions) Cd14 apc alexa fluor 750, by Thermo Fisher Scientific (1 mentions) Cd19 apc alexafluor750, by Thermo Fisher Scientific (1 mentions) Bv605 ccr6, by BioLegend (1 mentions) Cxcr3 pe cy7, by BD (1 mentions) Tnfα efluor 450, by Thermo Fisher Scientific (1 mentions) Ifnγ alexa fluor 700, by BD (1 mentions) Hla dr pe, by BD (1 mentions) Cytofix cytoperm buffer, by BD (1 mentions) Cd4 fitc, by BD (1 mentions) Cytofix cytoperm fixation permeabilization kit, by BD (1 mentions) Cd8 buv496, by BD (1 mentions) Phorbol myristate acetate (pma), by Thermo Fisher Scientific (1 mentions) Live dead stain, by Thermo Fisher Scientific (1 mentions) Cd20 apc cy7, by BD (1 mentions) Igm bv605, by BD (1 mentions)

11 protocols using cd3 bv650

Multiparameter Flow Cytometry of CSF Immune Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Flow cytometry was conducted using an LSRFortessa (BD Biosciences). A panel consisting of antibodies conjugated to six different fluorophores was used to classify subsets of memory T cells and for drop-seq. Antibodies used were: CD8α-Pacific blue (BioLegend), CD3-BV650 (BD Biosciences), CD45RA-APC-Cy7 (BioLegend), CCR7–488 (Bio-Legend), IL-7Rα-PE (BioLegend) and CD27-PE-Cy7 (BioLegend). For characterization of CSF cells, this same panel was used, but CD19-PE-Cy5 (BioLegend) and CD14-Qdot-705 (Thermo Fisher Scientific) were included. For sorting CSF T cells for TCR plate-seq, the following antibodies were used: CD8α-PE (BioLegend), CD161-PE-Cy7 (BioLegend), CXCR3-APC (BioLegend), CD4-APC-Alexa700 (Thermo Fisher Scientific), CD39-APC-Cy7 (BioLegend), CD38-FITC (BioLegend), PD-1-BV421 (BD Biosciences), CD45RA-BV605 (BD Biosciences), CD3-BV650 (BD Biosciences), CD27-BV786 (BD Biosciences) and CD127-BUV395 (BD Biosciences). For each experiment, a compensation matrix was developed using singly stained and unstained controls or fluorescent beads, and all analysis was conducted in Cytobank.

Gate D., Saligrama N., Leventhal O., Yang A.C., Unger M.S., Middeldorp J., Chen K., Lehallier B., Channappa D., De Los Santos M.B., McBride A., Pluvinage J., Elahi F., Tam G.K., Kim Y., Greicius M., Wagner A.D., Aigner L., Galasko D.R., Davis M.M, & Wyss-Coray T. (2020). Clonally expanded CD8 T cells patrol the cerebrospinal fluid in Alzheimer’s disease. Nature, 577(7790), 399-404.

+ Open protocol

+ Expand

Multi-parametric Phenotyping of Immune Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

After stimulation, cells were washed, stained with LIVE/DEAD^® Fixable Near-IR Stain (Invitrogen) and, subsequently, surface stained with the following antibodies: CD14-APC/Alexa Fluor 750 (Invitrogen) and CD19-APC/Alexa Fluor 750 (Invitrogen), CD4-FITC (BD), CD27-BV711 (BD), CCR4-BV510 (Biolegend), CCR6-BV605 (Biolegend), CXCR3-PE-Cy7 (BD Biosciences), KLRG1-PerCP/eFluor 710 (eBioscience), and HLA-DR-PE (BD). Cells were then fixed and permeabilized using Cytofix/Cytoperm buffer (BD) and stained with CD3-BV650 (BD), IL-2-PE/Dazzle™ (Biolegend), TNFα-eFluor 450 (eBioscience), and IFNγ-Alexa Fluor 700 (BD). Finally, cells were washed and fixed in 1% formaldehyde in PBS. Samples were acquired on a LSR-II (BD) and analyses were performed using FlowJo (Treestar). A positive IFNγ response was defined as at least twice the background measured in the presence of co-stimulatory antibodies without antigen. Cell polyfunctionality was analyzed using Pestle and Spice software (19 (link)). The gating strategy is presented in Figure S1 in Supplementary Material.

Riou C., Berkowitz N., Goliath R., Burgers W.A, & Wilkinson R.J. (2017). Analysis of the Phenotype of Mycobacterium tuberculosis-Specific CD4+ T Cells to Discriminate Latent from Active Tuberculosis in HIV-Uninfected and HIV-Infected Individuals. Frontiers in Immunology, 8, 968.

+ Open protocol

+ Expand

Quantifying IL-9 and IL-33 T Cells in Rhesus PBMCs

Check if the same lab product or an alternative is used in the 5 most similar protocols

To measure IL-9 and IL-33 expressing T cells, Rhesus PBMCs were stimulated with 50 ng/ml of PMA (cat # 1201/1, Fisher Scientific) and 2.5 μM of Ionomycin in DMEM medium and incubated for 4 h in the presence of GolgiStop (BD-Biosciences) at 37°C. Cells were washed at 350 g for 5 min with MACS buffer (MACS buffer+1% BSA). Cells were stained with live dead stain (cat # L34961, Molecular Probes) and incubated for 20 mins at room temperature. Cells were washed and stained with cell surface markers (CD4-BB790 (BD-custom), CD19-AF700(Beckman Coulter- cat # A78837), CD20-APC-Cy7(BD Biosciences cat # 335794), IgM-BV605 (BD-Biosciences- cat # 562977, CD3-BV650 (BD Biosciences cat # 563916), CD8-BUV496-cat # 564805), incubated for 1 hr. at 4°C. Cells were washed twice with MACS buffer and fixed/permeabilized according to manufacturer’s protocol (BD-Cytofix/Cytoperm Fixation/Permeabilization kit- Cat # 554714). After fixation/permeabilization cells were stained with IL-9-AF647 (BD Biosciences cat # 560813), incubated for 30 min at 4°C, washed twice in BD Perm/Wash buffer and analyzed by flow cytometry.

Sarkar S., Hessell A.J., Haigwood N.L, & Kobie J.J. (2020). Cloning and functional testing of rhesus macaque (Macaca mulatta) IL-9 and IL-33. Journal of medical primatology, 49(3), 144-152.

+ Open protocol

+ Expand

Multicolor Flow Cytometry for Treg Immunophenotyping

Check if the same lab product or an alternative is used in the 5 most similar protocols

Multicolour flow cytometry was used to assess the immunophenotype of Tregs. Antibodies against the following molecules were provided by: CD3 BV650 (BD Biosciences), CD8 BV450 (BD Biosciences), CD4 APC-H7 (BD Biosciences), CD25 PerCPCy5.5 (BD Biosciences), CD73 eFluor 450 (eBioscience™), PD-1 BV 650 (Biolegend). Dead cells were stained by LIVE/DEAD^® Fixable Blue Dead Cell Stain Kit (Life Technology). For intracellular staining, cells were fixed and permeabilized using the FoxP3/Transcription Factor Staining Buffer Set (eBioscience), and then stained with FoxP3 Alexa Fluor 488 (eBioscience), IL13-PE (eBioscience) and Granzyme B APC (BioLegend). Appropriate isotype controls were used to set the quadrants and to evaluate background staining. Cells were analysed using an LSRII flow cytometer with BD FACSDIVA software.

Faridar A., Thome A.D., Zhao W., Thonhoff J.R., Beers D.R., Pascual B., Masdeu J.C, & Appel S.H. (2020). Restoring regulatory T-cell dysfunction in Alzheimer’s disease through ex vivo expansion. Brain Communications, 2(2), fcaa112.

+ Open protocol

+ Expand

Immunoprofiling of Tumor Microenvironment

Check if the same lab product or an alternative is used in the 5 most similar protocols

Antibodies for immunohistochemical staining: anti-CD8α (D4W2Z, 98941), Foxp3 (D6O8R, 12653S), F4/80 (D2S9R, 70076S), CD206 (E6T5J, 24595S) were obtained from Cell Signaling Technology (Danvers, MA). TUNEL Reagent Test Kit (11684795910) was purchased from Roche (Shanghai, China).
Antibodies for Flow Analysis: CD8α-PerCP-Cy5.5 (53–6.7, 561109), CD25-BV510 (PC61, 563037), CD279 (PD-1)-BV605 (RMP1–30, 748267), CD3-BV650 (145-2C11, 564378), IL-10-BV711 (JES5-16E3, 564081), IL-17A-BV786 (TC11-18H10, 564171), CD45-BUV395 (30-F11, 565967), CD366 (Tim-3)-PE (5D12, 566346), STAT3-PE-CF594 (4/P-STAT3, 562673), CD4-PE-Cy7 (GK1.5, 563933), CD206-Alexa Fluor® 647 (MR5D3,565250), F4/80-BV510 (T45-2342,743280) were all obtained from BD Bioscience (San Jose, CA, USA). Foxp3-Alexa Fluor® 647 (150D, 320014), TGF-β1-FITC (TW7-16B4, 141414), Granzyme B-BV421(QA18A28, 396414) were purchased from BioLegend (San Diego, CA, USA). Fixable viability Dye eFluor^TM 780 (65-0865-14) were purchased from eBioscience^TM (San Diego, CA, USA).

Huang L., Zhao Y., Shan M., Wang S., Chen J., Liu Z, & Xu Q. (2023). Targeting crosstalk of STAT3 between tumor-associated M2 macrophages and Tregs in colorectal cancer. Cancer Biology & Therapy, 24(1), 2226418.

+ Open protocol

+ Expand

Modulating T Cell Activation in CLL

Check if the same lab product or an alternative is used in the 5 most similar protocols

T cells were magnetically purified from PBMC using a PAN T-cell isolation kit (Miltenyi Biotec) from healthy donors. Healthy donor T cells (1 × 10⁵) were then cultured with bead-purified (Miltenyi Biotec) CD19+ B cells from CLL patients (1 × 10⁶) at an optimal 1:10 ratio per well in the presence or absence of CXCL12 (250 ng/ml), lenalidomide (10 µM), CXCR4, or mouse anti-human IL-10 blocking antibodies (2 µg/ml, R&D). Each experiment was performed in duplicate. After 48 h of culture, T cells were magnetically purified again as described above (purity >95%) and activated with anti-CD3/CD28 magnetic Dynabeads (Invitrogen) for 6 h. A negative control (no stimulation) was included in every experiment. Brefeldin A (10 µg/mL) and CD107a PE-CF594 (BD Biosciences) were added to the culture. Cells were stained with a Live/Dead-Aqua (Invitrogen), CD3-BV650, CD8-FITC, CD4-APC-Cy7, fixed/permeabilized (BD Biosciences) followed by intracellular staining with IFN-γ-V450, TNF-α-Alexa 700, and IL-2-PE (BD Biosciences).

Shaim H., Estrov Z., Harris D., Hernandez Sanabria M., Liu Z., Ruvolo P., Thompson P.A., Ferrajoli A., Daher M., Burger J., Muftuoglu M., Imahashi N., Li L., Liu E., Alsuliman A.S., Basar R., Nassif Kerbauy L., Sobieski C., Gokdemir E., Kondo K., Wierda W., Keating M., Shpall E.J, & Rezvani K. (2018). The CXCR4–STAT3–IL-10 Pathway Controls the Immunoregulatory Function of Chronic Lymphocytic Leukemia and Is Modulated by Lenalidomide. Frontiers in Immunology, 8, 1773.

+ Open protocol

+ Expand

Multiparameter Flow Cytometry Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Multiparameter FACS analysis was performed using a monoclonal antibody (mAb) panel to measure activation in blood [18] . The following mAb combinations, relevant for this manuscript, were used: CD45^BUV496 (Becton & Dickinson, Franklin Lakes, NJ, USA (BD), cat 741185), HLA-DR^BUV805 (BD, cat 748338), CD20^V450 (BD, cat 655872), CD16^BV605 (BD, cat 563916), CD3^BV650 (BD, 563916), CD40^BV750 (BD, cat 746948), CD14^PE-TxRed (Beckman Coulter, cat B92391), CD123^PerCP-Cy5.5 (BD cat 558714), CD86^Alexa647 (Biolegend, San Diego, CA, USA, cat 305416), CD1c^APC (Ebioscience cat 17-0015-42). Acquisition was performed on an Aurora flow cytometer (Cytek, Fremont, Ca, USA) and analyzed using FlowJo software. For analysis, the CD45 positive cells with a low side scatter (SSC) profile were first gated and then singlets were selected using a forward scatter (FSC) height versus area plot. Then, HLA-DR positive/CD3, CD20 and CD14 negative cells were selected and CD123 was plotted against CD1c to identify the plasmacytoid DC (pDC)(CD123 positive/CD1c negative) and classical DC2 (cDC2)(CD123 negative/CD1c positive) populations. Expression of CD40 and CD86 was quantified as the mean geometric mean of the fluorescence intensity (MFI).

Meijer L., Böszörményi K.P., Bakker J., Koopman G., Mooij P., Verel D., Fagrouch Z., Verstrepen B.E., Funke U., Mooijer M.P., Langermans J.A., Verschoor E.J., Windhorst A.D, & Stammes M.A. (2022). Novel application of [18F]DPA714 for visualizing the pulmonary inflammation process of SARS-CoV-2-infection in rhesus monkeys (Macaca mulatta). Nuclear Medicine and Biology, 112-113, 1-8.

+ Open protocol

+ Expand

Isolation and Profiling of Human Skin T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Primary human T cells (n = 52) were isolated by digestion of fresh human skin biopsies (Ø 6 mm) in RPMI containing FCS, Collagenase type IV (Worthington), and Deoxyribonuclease I (Sigma) at 37 °C overnight followed by dissociation using the gentleMACS Dissociator (Miltenyi Biotec). Freshly isolated skin T cells were passed over a cell strainer and directly used for flow cytometric analysis. For flow cytometric analysis, T cells were stimulated with PMA/Ionomycin (10 ng/ml and 1 µg/ml, respectively) (both Sigma) for 5 h in the presence of Brefeldin A and Monensin (both BD Biosciences). Surface staining was performed at 4 °C and followed by fixation/ permeabilization using the fixation/permeabilization kit (BD Biosciences). Staining of intracellular cytokines was performed at room temperature. Antibodies used were CD3-Bv650 (#563852, clone UCHT1, dilution 1:50), CD4-BV421 (#562842, clone L200, dilution 1:20), CD8-APCCy7 (#557834, clone SK1, dilution 1:20) (BD Biosciences), IL-17A-PeCy7(#512315, clone BL468, dilution 1:20), IFN-γ-PerCPCy5.5 (#506528, clone B27, dilution 1:100), TNF-α-BV510 (#502950, clone MAB11, dilution 1:100) (BioLegend), IL-22-Pe (#12-7229-41, clone 22URTI, dilution 1:20, eBioscience), IL-10-APC (#130-108-135, clone JES3-9D, dilution 1:10, Miltenyi Biotec). Flow cytometry data was analysed and visualized using FlowJo 10.7.1.

Schäbitz A., Hillig C., Mubarak M., Jargosch M., Farnoud A., Scala E., Kurzen N., Pilz A.C., Bhalla N., Thomas J., Stahle M., Biedermann T., Schmidt-Weber C.B., Theis F., Garzorz-Stark N., Eyerich K., Menden M.P, & Eyerich S. (2022). Spatial transcriptomics landscape of lesions from non-communicable inflammatory skin diseases. Nature Communications, 13, 7729.

+ Open protocol

+ Expand

Multiparameter Flow Cytometry Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cell staining was performed using various combinations of the following Abs: CD3-BV650 (BD, clone UCHT1), CD4-BV786 (BD, clone SK3), CD8-APC-H7 (BD, clone SK1), CD103-BV605 (BD, clone Ber-ACT8), CD69-BV421 (BD, clone FN50), CD19-APC-H7 (BD, clone HIB19), CD24-PE (BD, clone ML5), CD27-PE-Cy7 (BD, clone M-T271), CD38-PerCP-Cy5.5 (BD, clone HIT2), IgD-FITC (BD, clone IA6-2), CXCL13-APC (Invitrogen, clone 53,610). Surface staining was performed for 30 min at 4 °C. Intracellular staining was performed by the use of Intrasure kit (BD) according with the manufacturer instructions. Cells were immediately acquired in the BD FACSCelesta flow cytometer and analyzed by the use of BD FACSDiva software (RRID:SCR_001456).

Di Modugno F., Di Carlo A., Spada S., Palermo B., D'Ambrosio L., D'Andrea D., Morello G., Belmonte B., Sperduti I., Balzano V., Gallo E., Melchionna R., Panetta M., Campo G., De Nicola F., Goeman F., Antoniani B., Carpano S., Frigè G., Warren S., Gallina F., Lambrechts D., Xiong J., Vincent B.G., Wheeler N., Bortone D.S., Cappuzzo F., Facciolo F., Tripodo C., Visca P, & Nisticò P. (2024). Tumoral and stromal hMENA isoforms impact tertiary lymphoid structure localization in lung cancer and predict immune checkpoint blockade response in patients with cancer. eBioMedicine, 101, 105003.

+ Open protocol

+ Expand

Multiparametric Analysis of SARS-CoV-2 Antibody Profiles

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cryopreserved PBMCs were thawed and washed twice with 10 ml of fluorescence-activated cell sorting (FACS) buffer (1× PBS containing 2% FBS and 1 mM EDTA) and resuspended in 100 μl of PBS containing Zombie UV LIVE/DEAD dye at 1:200 dilution (BioLegend, #423108) and incubated at room temperature for 15 min. After washing, cells were incubated with an antibody cocktail for 1 hour protected from light on ice. The following antibodies were used: IgD phycoerythrin (PE; Southern Biotech, #2030–09), IgM peridinin chlorophyll protein–Cy5.5 (BioLegend, #314512), CD20 allophycocyanin-H7 (BD Biosciences, #560734), CD27 PE-Cy7 (BioLegend, #302838), CD14 brilliant violet (BV) 650 (BioLegend, #301836), CD16 BV650 (BioLegend, #302042), IgG brilliant UV 395 (BD Biosciences, #564229), CD3 BV650 (BD Biosciences, #563916), CD21 PE-CF594 (BD Biosciences, #563474), Alexa Fluor 488–labeled Beta Spike protein (antibodies-online, #ABIN6963740), Alexa Fluor 647– labeled Omicron Spike protein (Sino Biological, #40589-V08H26), and BV421-l abeled Wuhan Spike protein (Sino Biological, #40589-V27B-B). All antibodies were used as per the manufacturer’s instruction, and the final concentration of each probe was 0.1 μg/ml. Cells were washed twice in FACS buffer and immediately acquired on a BD FACSAria III. FlowJo software v10 (TreeStar Inc.) was used for data analysis.

Arunachalam P.S., Feng Y., Ashraf U., Hu M., Walls A.C., Edara V.V., Zarnitsyna V.I., Aye P.P., Golden N., Miranda M.C., Green K.W., Threeton B.M., Maness N.J., Beddingfield B.J., Bohm R.P., Scheuermann S.E., Goff K., Dufour J., Russell-Lodrigue K., Kepl E., Fiala B., Wrenn S., Ravichandran R., Ellis D., Carter L., Rogers K., Shirreff L.M., Ferrell D.E., Deb Adhikary N.R., Fontenot J., Hammond H.L., Frieman M., Grifoni A., Sette A., O’Hagan D.T., Van Der Most R., Rappuoli R., Villinger F., Kleanthous H., Rappaport J., Suthar M.S., Veesler D., Wang T.T., King N.P, & Pulendran B. (2022). Durable protection against the SARS-CoV-2 Omicron variant is induced by an adjuvanted subunit vaccine. Science translational medicine, 14(658), eabq4130.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!