B220 pacific blue

Manufactured by BD

Sourced in United States

The B220-Pacific Blue is a lab equipment product. It is a fluorescent dye used for flow cytometry analysis. The B220-Pacific Blue emits a blue fluorescent signal when excited by a 405 nm laser.

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Lab products found in correlation

Cd43 apc, by BD (3 mentions) Cd24 pe cy7, by BD (3 mentions) Bp1 pe, by Thermo Fisher Scientific (3 mentions) Cd43 fitc, by BD (2 mentions) Total h3, by Cell Signaling Technology (2 mentions) α tubulin, by Merck Group (2 mentions) Cd45.1 pecy7, by Thermo Fisher Scientific (2 mentions) Hmgn1, by Aviva Systems Biology (2 mentions) Hmgn1, by Abcam (2 mentions) Bp1 fitc, by Thermo Fisher Scientific (2 mentions) Cd45.2 apc, by Thermo Fisher Scientific (2 mentions) H3k27me3, by Cell Signaling Technology (2 mentions) H3k4me3, by Abcam (2 mentions) H3k27ac, by Abcam (2 mentions) Cd19 apc, by BioLegend (2 mentions) Cd3 fitc, by BD (2 mentions) Igd apc cy7, by BD (2 mentions) Cd21 fitc, by BD (2 mentions) Cd23 pe, by BD (2 mentions) Cd138 pe, by BD (2 mentions)

9 protocols using b220 pacific blue

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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Western Blotting and Flow Cytometry Antibodies

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Western blotting antibodies were HMGN1 (Aviva Systems Biology, #ARP38532_P050), HMGN1 (Abcam, #ab5212), mouse HMGN1 (affinity purified rabbit polyclonal)^{34 (link),35 (link)}, H3K27me3 (Cell Signaling Technologies, #9733, rabbit polyclonal), total H3 (Cell Signaling Technologies, #9715, rabbit polyclonal), and α-tubulin (Sigma, #T9026, mouse monoclonal). Flow cytometry antibodies were B220-Pacific Blue (BD Pharmingen, #558108, clone RA3-6B2), CD43-APC (BD, #560663, clone S7) or CD43-FITC (BD, #561856, clone S7), CD24-PE-Cy7 (BD, #560536, clone M1/69), BP1-PE (eBiosciences, 12-5891, clone 6C3) or BP1-FITC (eBiosciences, 11-5891, clone 6C3), CD45.1-PE-Cy7 (eBiosciences, 25-0453, clone A20), and CD45.2-APC (eBiosciences, 17-0454, clone 104). ChIP-seq antibodies were H3K27me3 (Cell Signaling Technologies, #9733), H3K4me3 (Abcam, #ab8580), and H3K27ac (Abcam, #ab4729).

Lane A.A., Chapuy B., Lin C.Y., Tivey T., Li H., Townsend E.C., van Bodegom D., Day T.A., Wu S.C., Liu H., Yoda A., Alexe G., Schinzel A.C., Sullivan T.J., Malinge S., Taylor J.E., Stegmaier K., Jaffe J.D., Bustin M., te Kronnie G., Izraeli S., Harris M., Stevenson K.E., Neuberg D., Silverman L.B., Sallan S.E., Bradner J.E., Hahn W.C., Crispino J.D., Pellman D, & Weinstock D.M. (2014). Triplication of a 21q22 region contributes to B cell transformation through HMGN1 overexpression and loss of histone H3 lysine 27 trimethylation. Nature genetics, 46(6), 618-623.

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Comprehensive Hematopoietic Profiling in Mice

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Peripheral blood from each mouse was subjected to complete blood count. Tibias, femurs, iliac crests, spines, ulnae, radii, and humeri were harvested for bone marrow cells. Spleen and thymus were collected for lymphocyte staining. Flow cytometry staining for all hematopoietic subpopulations including T cell developmental stages was performed as previously described (Yu et al., 2015a (link)). For B cell development, the following scheme was used: B220-Pacific Blue (BD Biosciences 558108), IgM-PE-Cy5 (eBiosciences 15-5790-82), CD43-fluorescein isothiocyanate (FITC) (eBiosciences 11-0431-82), CD24-APC (Biolegend 101814), and BP1-PE (eBiosciences 12-5891-83). Definitions of stages of B cell maturation were as follows: A′ pre-pro-B (IgM⁻B220⁺CD43⁺BP1⁻CD24⁻), B′ pro-B (IgM⁻B220⁺CD43⁺BP1⁻CD24⁺), C′ pro-B (IgM⁻B220⁺CD43⁺BP1⁺CD24^lo), C″ pro-B (IgM⁻B220⁺CD43⁺BP1⁺CD24^hi), pro-B (IgM⁻B220⁺CD43⁺), pre-B (IgM⁻B220⁺CD43⁻), B progenitors (IgM⁻B220⁺), immature B (IgM⁺B220^lo), and mature B (IgM⁺B220^hi). For cell-cycle analysis, bromodeoxyuridine-FITC and 7AAD, or Ki67-FITC and DAPI staining were coupled with staining of specific populations to reveal their cell-cycle status.

Yu V.W., Lymperi S., Oki T., Jones A., Swiatek P., Vasic R., Ferraro F, & Scadden D.T. (2016). Distinctive Mesenchymal-Parenchymal Cell Pairings Govern B Cell Differentiation in the Bone Marrow. Stem Cell Reports, 7(2), 220-235.

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Western Blotting and Flow Cytometry Antibodies

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Tumor-Draining Lymph Node Cell Profiling

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Tumor tissue and axial and brachial TDLNs were harvested at indicated time points, and then, single cell suspensions were generated. B cell cultures were harvested at 72 hours following plating and washed twice with PBS. Cells were stained with the following fluorochrome conjugated antibodies for surface markers: CD11b-BV421 (BD Biosciences, San Jose, CA, clone M1/70), Gr1-FITC (BD Biosciences, San Jose, CA, clone RB6-8C5), CD19-APC (Biolegend, San Diego, CA, clone 6D5), and B220-Pacific Blue (BD Biosciences, San Jose, CA, clone RA3-6B2). Additionally, the following antibodies were used to stain for intracellular markers: IL-17A-PE (BD Biosciences, San Jose, CA, clone TC11-18H10), RORγt-APC (BD Biosciences, San Jose, CA, clone Q31-378), and COX2-Ax488 (Santa Cruz Biotechnology, Dallas, TX, clone H-3). We used a FOXP3 Fixation/Permeabilization Kit from Invitrogen (Carlsbad, CA) for the above staining procedure. A BD LSRII Flow Cytometer System was used for the flow cytometric analyses. Data were acquired from 200,000 TDLN cells by using the BD-LSRII instrument, and these data were analyzed by using FlowJo software. Positive gates (gates containing antibody binding cells) were established using fluorescence minus one (FMO) controls (15 (link)). FMO controls are the experimental cells stained with all the fluorophores minus one.

Falk-Mahapatra R, & Gollnick S.O. (2022). Photodynamic Therapy-induced Cyclooxygenase 2 Expression in Tumor-Draining Lymph Nodes Regulates B Cell Expression of Interleukin 17 and Neutrophil Infiltration. Photochemistry and photobiology, 98(5), 1207-1214.

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Analysis of B Cell Differentiation

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For analysis of B cell differentiation, 5 x 10⁴ transduced HSCs were transferred to flasks of OP9 stromal cells and co-cultured in B lymphoid-promoting medium (Alpha MEM supplemented with 20% FBS, 1% penicillin-streptomycin, 50 μM 2-mercaptoethanol, 10 ng/mL SCF, 10 ng/mL Flt3L and 10 ng/mL mIL-7 (PeproTech, Rocky Hill, NJ), and 250 ng/mL amphotericin B). Cells were harvested by trypsinization and resuspended in sorting buffer after 7 days of co-culture. Hardy fraction analysis of B cell differentiation was performed as previously described [35 (link)]. Briefly, cells were stained with 7-AAD (eBioscience) and the following antibodies: B220-Pacific blue, CD43-APC, BP-1-BV605, CD24-PE-Cy7, IgD-APC-Cy7, and IgM-PE-CF594 (BD Biosciences). Single-color stained cells and UltraComp beads (eBioscience) were used for compensation and to establish Hardy fraction gating. The following controls were used to exclude OP9 cells and non-transduced cells from ZsGreen⁺/RFP⁺ analysis: OP9 cells alone, non-transduced HSPCs alone, and OP9 cells with non-transduced HSPCs. Samples were analyzed on an LSR II flow cytometer and FACS plots were generated using FlowJo software.

Gant V.U., Junco J.J., Terrell M., Rashid R, & Rabin K.R. (2021). Enhancer polymorphisms at the IKZF1 susceptibility locus for acute lymphoblastic leukemia impact B-cell proliferation and differentiation in both Down syndrome and non-Down syndrome genetic backgrounds. PLoS ONE, 16(1), e0244863.

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Comprehensive B Cell Immunophenotyping

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Cells (1× 10⁶) were collected by centrifugation and washed twice in PBS/2% FCS, and the cell pellet was resuspended in 100 μl of PBS/2% FCS. Fluorophore-conjugated surface marker Abs were added at a 1:100 dilution and incubated on ice for 20 min. Cells were washed in PBS/2% FCS and resuspended in PBS/2% FCS for analysis on a BD LSR II Flow Cytometer. The Abs used in the FACS analysis were as follows: CD21-FITC (553818) BD Biosciences Rat, CD23-PE (561773) BD Pharmingen Rat, CD23-Pacific blue (101616) BioLegend Rat, IgD-allophycocyanin (405713) BioLegend Rat, IgM-PE-Cy7 (406513) BioLegend Rat, B220-Pacific blue (558108) BD Biosciences Rat, B220-PE (553089) BD Biosciences Rat, and CD22-PE (BD 553384). The LIVE/DEAD Fixable Aqua Dead Cell Stain Kit (Thermo Fisher Scientific) was used to distinguish between live and dead cells. Data were analyzed with FlowJo software. The gating strategy was that cells were initially gated for live lymphocytes followed by analysis of B220⁺ cells.

Thomsen I., Kunowska N., de Souza R., Moody A.M., Crawford G., Wang Y.F., Khadayate S., Whilding C., Strid J., Karimi M.M., Barr A.R., Dillon N, & Sabbattini P. (2021). RUNX1 Regulates a Transcription Program That Affects the Dynamics of Cell Cycle Entry of Naive Resting B Cells. The Journal of Immunology Author Choice, 207(12), 2976-2991.

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Multiparametric Immune Cell Analysis

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Single immune cell suspensions were prepared from spleen or peritoneal lavage after red blood cell lysis. Cells were counted on a BD Fortessa Flow cytometer using ‘Fluoresbrite ^TM Calibration Grade 6.0 micron YG microspheres (Polysciences). Cells were incubated with anti-CD16/CD32 (Fc block, clone 2.4G2; BD Bioscience) plus 2% normal mouse and normal rat serum and stained with various combinations of the following Abs in FACS buffer (PBS containing 2% FBS and 2 mM EDTA) for 30 min at 4°C. Murine reactive antibodies, including B220-Pacific Blue, CD3-PerCPCy5.5, CD4-Pacific Blue, CD5-PerCPCy5.5, CD8-APC Cy, CD11b APC Cy7, CD21 FITC, CD23 PE, CD38-FITC, CD95 PE Cy7, CD138-PE, IgD APC Cy7, IgM FITC, GL-7 biotin, Streptavidin BV500, Ki67-APC, Ly6G PE Cy7, and Ly6C PerCP Cy5.5 were all purchased from BD Bioscience.

Katewa A., Suto E., Hui J., Heredia J., Liang J., Hackney J., Anderson K., Alcantar T.M., Bacarro N., Dunlap D., Eastham J., Paler-Martinez A., Rairdan X.Y., Modrusan Z., Lee W.P., Austin C.D., Lafkas D, & Ghilardi N. (2021). The peptide symporter SLC15a4 is essential for the development of systemic lupus erythematosus in murine models. PLoS ONE, 16(1), e0244439.

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Multiparameter Flow Cytometry of Lymphocytes

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Lymphocytes from local lymph nodes were collected and stained with different fluorescence-labeled antibody cocktails as we previously described [27 (link)] to detect the percentages of CD4⁺ T cells (CD3-FITC, CD4-PE), CD8⁺ T cells (CD3-FITC, CD8a-APC), IFN-γ secreting cells (CD3-FITC, CD8-APC, IFN-γ-PE-Cy5-5), Tfh (T follicular helper) cells (CD45-APC-Cy7 (BD Pharmingen, San Diego, CA, USA), CD4-FITC (BD Pharmingen, USA), CD185-APC, PD-1-Pacific Blue (BD Pharmingen, USA)), germinal center (GC) B cells (B220-APC-Cy7, CD45-APC-Cy7 (BD Pharmingen, USA), CD95-PE, GL-7-APC), and plasma cells (B220-Pacific Blue, CD27-PE-Cy7, CD138-PE). Except where indicated, the remaining antibodies were purchased from BioLegend, San Diego, CA, USA. The stained cells were assessed by flow cytometry (BD LSRFortessa ^TM Cell Analyzer, San Jose, CA, USA), followed by FlowJo software analysis.

Meng Z., Ma D., Duan S., Zhang J., Yue R., Li X., Gao Y., Li X., Zeng F., Xu X., Jiang G., Liao Y., Fan S., Niu Z., Li D., Yu L., Zhao H., Xu X., Wang L., Zhang Y., Liu L, & Li Q. (2022). Immunological Study of Combined Administration of SARS-CoV-2 DNA Vaccine and Inactivated Vaccine. Vaccines, 10(6), 929.

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