Cd19 apc

Manufactured by BioLegend

Sourced in United States

CD19-APC is a fluorescent-conjugated antibody that binds to the CD19 antigen, which is expressed on B cells. It is designed for use in flow cytometry applications to identify and quantify B cells in biological samples.

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CD19-APC-Cy7 (45 citations) Anti-CD19-APC-Cy7 (17 citations) APC anti-mouse CD19 (10 citations) α-CD19 APC (6 citations) Mouse anti-human CD19-APC (3 citations) APC-conjugated anti-CD19 (3 citations) APC-anti-human CD19 (clone HIB19, (3 citations) Anti-CD19-APC (6D5, (2 citations) α-human CD19-APC (HIB19; (2 citations) APC-conjugated CD19 (2 citations)

40 protocols using cd19 apc

Multiparametric Flow Cytometry Profiling of Hematopoietic Lineages

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Blood lymphoid. CD4-Fitc (BioLegend), CD8-PE-Cy7 (BD), B220–Pacific blue (BD), CD11b-PerCpCy5,5 (BD), CD3-APC (BioLegend); Blood myeloid and erythroid: CD3-Fitc (BD), CD19-Fitc (BD), B220-Fitc (BD), Gr1-PE (BD), F4/80-Pacific blue (BioLegend), CD11b-APC (BD);
BM lymphoid. IgD-Fitc (BD), CD25-PE (BioLegend), CD11b-PerCpCy5,5 (BD), IgM-PECy7 (Southern Biotech), B220-Pacific blue (BD), cKit-APC (BD); BM myeloid: Gr1-PE (BD), CD11b-PerCpCy5,5 (BD), Ter119-PECy7 (BD), F4/80–Pacific blue (BioLegend), CD19-APC (BioLegend); BM megakaryocyte: CD3-Fitc (BD), CD41-PE (BioLegend), CD11b-PerCpCy5,5 (BD), B220–Pacific blue (BD), CD19-APC (BioLegend); HSPC and CLP: Strep-APC/CY7 (Southern Biotech), cKit-APC (BD), SCA1-PacBlue (BioLegend), CD34-FITC (eBioscience), CD135-PE (BioLegend), CD150-PECy7 (BioLegend), CD127-PErCpCy5,5 (BioLegend); Hematopoietic progenitors: Strep-APC/CY7 (Southern Biotech), cKit-APC (BD), SCA1-PacBlue (BioLegend), CD34-FITC (eBioscience), CD16/32-PerCpCy5,5 (eBioscience), CD127-PE (eBioscience).

Alendar A., Lambooij J.P., Bhaskaran R., Lancini C., Song J.Y., van Vugt H., Snoek M, & Berns A. (2020). Gene expression regulation by the Chromodomain helicase DNA-binding protein 9 (CHD9) chromatin remodeler is dispensable for murine development. PLoS ONE, 15(5), e0233394.

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Quantifying Somatic Hypermutation in Murine B Cells

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Single-cell suspensions from Peyer’s patches of 30-31 weeks old mice were first incubated with anti-mouse CD16/32 (BioLegend) and then labeled with B220-FITC– (BioLegend), CD19-APC– (BioLegend), CD38-Alexa700– (Thermo Scientific), and CD95/Fas-PE– (BD-Biosciences) conjugated antibodies. Non-germinal center (CD38⁺ Fas⁻) and germinal center B cells (CD38⁻ Fas⁺) were sorted on an Aria BD sorter. Genomic DNA was extracted and the 5′ portions of J_H4 (Igh) and J_K5 (Igk) introns were amplified by PCR using Phusion High-Fidelity DNA Polymerase (Thermo Scientific). The 800 bp J_H4 and 700 bp J_K5 PCR products were gel extracted, cloned into a pCR2.1 vector using the TOPO TA Cloning Kit (Invitrogen) and sequenced. Mutations were quantified over 510 bp downstream J_H4 and 536 bp downstream J_K5 gene segments. Primers used for SHM analysis (Rouaud et al., 2013 (link), Sander et al., 2015 (link)) are listed in Table S5.

Delgado-Benito V., Rosen D.B., Wang Q., Gazumyan A., Pai J.A., Oliveira T.Y., Sundaravinayagam D., Zhang W., Andreani M., Keller L., Kieffer-Kwon K.R., Pękowska A., Jung S., Driesner M., Subbotin R.I., Casellas R., Chait B.T., Nussenzweig M.C, & Di Virgilio M. (2018). The Chromatin Reader ZMYND8 Regulates Igh Enhancers to Promote Immunoglobulin Class Switch Recombination. Molecular Cell, 72(4), 636-649.e8.

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Comprehensive Immune Cell Profiling in BAL

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Absolute cell counts in BAL were determined by flow cytometry with Precision Count Beads (BioLegend) by staining 100 μL of BAL fluid with pre-titrated amounts of CD45 FITC (clone HI30), CD14 PerCP-Cy5.5 (clone M5E2), CD3 PE-Cy7 (clone UCHT1), CD8 BV421 (clone RPA-T8), CD4 APC-Cy7 (clone OKT4) and CD19 APC (clone HIB19). 1 mL of FACS Lysing Solution (BD Biosciences) was added to each sample. 100 μL of Precision Count Beads were added immediately prior to flow cytometry to allow quantification of absolute volume analyzed on the cytometer.
In a separate experiment, fresh BAL cells were analyzed using a panel of antibodies directed against the following antigens: CD3 FITC (clone UCHT1), CD4 APC-Cy7 (clone OKT4), CD8 BV421 (clone RPA-T8), CD14 APC (clone M5E2), CD16 BV570 (clone 3G8), CD19 BV750 (clone HIB19), CD20 Pacific Blue (clone 2H7), CD38 PE-Cy7 (clone HIT2), CD45 Alexa Fluor 532 (clone HI30), CD56 PE/Dazzle 594 (clone HCD56), and HLA-DR BV605 (clone L243). 500,000–1,000,000 fresh BAL cells were stained in BD Brilliant Buffer (BD Biosciences) with Zombie NIR Fixable Viability Marker (BioLegend). Samples were run on a Cytek Aurora spectral flow cytometer using SpectroFlo software (version 2, Cytek) before final analysis in FlowJo software (version 10, BD Biosciences).

Reynolds D., Guillamet C.V., Day A., Borcherding N., Guillamet R.V., Choreño-Parra J.A., House S.L., O’Halloran J.A., Zúñiga J., Ellebedy A.H., Byers D.E, & Mudd P.A. (2021). Comprehensive immunologic evaluation of bronchoalveolar lavage samples from human patients with moderate and severe seasonal influenza and severe COVID-19. Journal of immunology (Baltimore, Md. : 1950), 207(5), 1229-1238.

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Characterization of Activated T Cells in RA

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Previously frozen PBMCs from RA subjects were cultured at 5 × 10⁶ total cells/well within a 24 well plate in RPMI-1640 + 10% pooled human serum with 10 µg/ml of peptide. IL-2 (Novartis) was added at 325 IU/ml on day 6. After 14 days cells were stained for expression of CD25 FITC (BD), CD4 PerCP (BioLegend), Annexin V APC (BD), CD14 APC (BioLegend), CD19 APC (BioLegend), and tetramer before being run on a FACSCalibur. The data was analyzed by FlowJo software version 9.6.2 (Tree star).

James E., Rieck M., Pieper J., Gebe J.A., Yue B.B., Tatum M., Peda M., Sandin C., Klareskog L., Malmström V, & Buckner J.H. (2014). Citrulline specific Th1 cells are increased in rheumatoid arthritis and their frequency is influenced by disease duration and therapy. Arthritis & rheumatology (Hoboken, N.J.), 66(7), 1712-1722.

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CFSE-Labeled PBMC Proliferation Assay

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PBMCs were incubated with 1.25 μl/ml CFSE (Invitrogen) at 37°C. After 10 min, the cells were washed twice at 4°C with 5 ml Roswell Park Memorial Institute (RPMI) medium containing 10% fetal bovine serum (FBS). Cells were then resuspended in 600 μl of RPMI/10% FBS and seeded into 96-well plates along with 5 μg/ml phytohemagglutinin (PHA), 10 μg/ml lectin from pokeweed mitogen (PWM), and the same volume of RPMI for 72 h. After staining with CD3-PerCP (clone: HIT3a, BioLegend), CD4-PE-Cy7 (clone: RPA-T4, BioLegend), CD8-PE (clone: RPA-T8, BioLegend), and CD19-APC (clone: HIB19, BioLegend) antibodies, cells were analyzed and examined by flow cytometry (8 (link)).

Luo X., Liu Q., Jiang J., Tang W., Ding Y., Zhou L., Yu J., Tang X., An Y, & Zhao X. (2021). Characterization of a Cohort of Patients With LIG4 Deficiency Reveals the Founder Effect of p.R278L, Unique to the Chinese Population. Frontiers in Immunology, 12, 695993.

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Multiparameter Single-Cell Profiling

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Samples were processed in batches of 6 or 8 for pooling in single-cell RNA sequencing runs. All cells were stained with a general panel: DAPI, CD3-APC (HIT3a), CD19-APC (HIB19), CD20-APC (2H7), CD56-APC (5.1H11), CD14-FITC (M5E2), CD16-AF700 (B73.1), CD45-PE-Cy7 (HI30), HLA-DR-PE (L243) (BioLegend). At the same time, 1 uL of cell hashing antibody (HTO) was added to each sample (BioLegend). Samples were run on a SH800 cell sorter (Sony) to obtain flow cytometry data and sort both live CD45+ cells and dendritic cells. For samples from subjects enrolled in the MGH ED, dendritic cells were enriched separately with a MACS human pan-DC enrichment kit (Miltenyi Biotec). For sorting MS1 cells, the following panel was used: DAPI, CD3-APC (HIT3a), CD19-APC (HIB19), CD20-APC (2H7), CD56-APC (5.1H11), CD14-FITC (M5E2), CD45-AF700 (HI30), HLA-DR-PE-Cy7 (L243) (BioLegend) and IL1R2-PE (34141, ThermoFisher Scientific).

Reyes M., Filbin M.R., Bhattacharyya R.P., Billman K., Eisenhaure T., Hung D.T., Levy B.D., Baron R.M., Blainey P.C., Goldberg M.B, & Hacohen N. (2020). An immune cell signature of bacterial sepsis. Nature medicine, 26(3), 333-340.

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Murine Immune Cell Isolation and Culture

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All reagents and chemicals were purchased from Sigma-Aldrich unless stated otherwise. DMEM with 1 g/l glucose and FBS-qualified MSC were purchased from PAN Biotech (PAN-Biotech GmbH, Aidenbach, Germany), FavorPrep™ Tri-RNA Reagent was from Favorgen (Favorgen Biotech Corp, Vienna, Austria), PacI Fast Digest enzyme, SYBR™ Green master mix, Dynabeads mouse T-Activator CD3/CD28 and Lipofectamine 3000 were from Thermo Fisher (Thermo Fisher Scientific, Waltham, MA, USA). Antibodies, anti-mouse Sca-1(PE), CD73(Cy7), CD45(PE), CD4(BV421), CD8a(APCfire), and CD19(APC) were from BioLegend (San Diego, CA, USA), and CFSE (5(6)-Carboxyfluorescein diacetate N-succinimidyl ester) were from Invitrogen (Fisher Scientific, part of Thermo Fisher Scientific, Waltham, MA, USA). Polybrene was purchased from Santa Cruz Biotechnology (sc 134220; Dallas, TX, USA), TransFast was from Promega (E2431, WI, USA), Viromer Red was from Cambridge Bioscience (VR-01LB-00, Cambridge, UK), the K2 Transfection System—DNA & RNA Transfection was from Biontex Laboratories GmbH (München, Germany).

Dumitrescu M., Vacaru A.M., Trusca V.G., Fenyo I.M., Ionita R, & Gafencu A.V. (2021). K2 Transfection System Boosts the Adenoviral Transduction of Murine Mesenchymal Stromal Cells. International Journal of Molecular Sciences, 22(2), 598.

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Multicolor Flow Cytometry Immunophenotyping

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Cell suspensions were labeled with anti-human monoclonal antibodies (mAb) targeting the following cell-surface markers: CD45-FITC, CD3-PE, CD4-Pe Cy7, CD8-BV421 and CD19-APC (all from Biolegend). Washing and reagent dilutions were done with FACS buffer (PBS containing 2% fetal calf serum and 0.05% sodium azide (NaN3). All acquisitions were performed on a Cyan ADP (Beckman Coulter) flow cytometer. Data were analysed with FlowJo software (Ashland, OR). Cellular debris and dead cells were excluded by their light-scattering characteristics.

Campos N., Myburgh R., Garcel A., Vautrin A., Lapasset L., Nadal E.S., Mahuteau-Betzer F., Najman R., Fornarelli P., Tantale K., Basyuk E., Séveno M., Venables J.P., Pau B., Bertrand E., Wainberg M.A., Speck R.F., Scherrer D, & Tazi J. (2015). Long lasting control of viral rebound with a new drug ABX464 targeting Rev – mediated viral RNA biogenesis. Retrovirology, 12, 30.

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Multiparametric Flow Cytometry of Murine and Human B-Cells

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Single‐cell suspensions were stained with a mixture of 5 to 6 fluorophores including and 7‐aminoactinomycin D (7‐AAD) for viability and acquired on the BD Verse flow cytometer with data analyzed using FlowJo software (BD Biosciences). Human B‐cell subsets were identified using the following antibodies: CD19‐APC, CD27‐BV510, CD38‐APC‐Cy7, CD24‐BV421, and IL‐10‐PE‐Cy7 (all from BioLegend). Mouse B‐cell subsets were identified using the following antibodies: B220‐PE, CD23‐APC, CD21‐PE‐Cy7, IgM‐BV421, CD138‐APC, CD86‐PE‐Cy7, CD69‐APC‐Cy7, and IL‐10‐APC‐Cy7 (all from BioLegend). Intracellular IL‐10 expression was detected as previously reported.³³ Immunophenotype of human and murine B‐cell subsets analyzed are listed in Supplementary Table 1.

Lee W., Wang L., Yen M., Hsu P., Lee Y., Liu K., Lin K., Su Y., Sytwu H, & Yen B.L. (2021). Resident vs nonresident multipotent mesenchymal stromal cell interactions with B lymphocytes result in disparate outcomes. Stem Cells Translational Medicine, 10(5), 711-724.

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Comprehensive Immune Profiling by Flow Cytometry

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For performing flow cytometry, we used fluorescent-labeled monoclonal antibodies to particular surface molecules: CD45-APC/Cy7, CD3-PerCP, CD3-PE/Cy7, CD4-APC, CD4-APC/Cy7, CD8-PE/Cy7, CD25-APC, CD127-PerCP/Cy 5.5, CD14-PE/Cy7, CD19-APC, CD16-PerCP, CD56-PE/Cy7 (all Biolegend, San Diego, CA, USA). CD25-PE and HLA-DR-APC/Cy7 antibodies (Biolegend, San Diego, CA, USA) were used for evaluating of the expression of activation markers. Flow cytometry analysis was performed on FACS Canto II (BD, Piscataway, NJ, USA) machine by using FACS Diva (BD, Piscataway, NJ, USA) software. We used non-stained controls, FMO controls, and isotypic controls.

Knauer N., Pashkina E., Aktanova A., Boeva O., Arkhipova V., Barkovskaya M., Meschaninova M., Karpus A., Majoral J.P., Kozlov V, & Apartsin E. (2022). Effects of Cationic Dendrimers and Their Complexes with microRNAs on Immunocompetent Cells. Pharmaceutics, 15(1), 148.

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