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B220 percp cy5

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The B220-PerCP-Cy5.5 is a fluorochrome-conjugated antibody designed for flow cytometry applications. It is used to detect the expression of the B220 cell surface antigen, which is commonly expressed on B lymphocytes. The PerCP-Cy5.5 fluorochrome provides a specific emission spectrum that can be detected by flow cytometers.

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Lab products found in correlation

Facscanto 2 system, by BD (2 mentions) Foxp3 staining kit, by Thermo Fisher Scientific (2 mentions) Cd8 percp cy5, by Thermo Fisher Scientific (2 mentions) Cd62l fitc, by Thermo Fisher Scientific (2 mentions) Gl 7 fitc, by Thermo Fisher Scientific (2 mentions) Cd95 pe, by Thermo Fisher Scientific (2 mentions) Gr 1 pecy7, by Thermo Fisher Scientific (2 mentions) Cd11c fitc, by Thermo Fisher Scientific (2 mentions) Cd4 v500, by BD (2 mentions) Cd3 alexa fluor 700, by Thermo Fisher Scientific (2 mentions) Cd8 efluor605, by Thermo Fisher Scientific (2 mentions) Biotinylated antibodies against cd3, by Thermo Fisher Scientific (2 mentions) B6 rag mice, by Jackson ImmunoResearch (2 mentions) Anti cd16 32 2.4g2 and normal mouse sera, by Jackson ImmunoResearch (2 mentions) Cd11b pe tr, by Thermo Fisher Scientific (2 mentions) Cd44 efluor450, by Thermo Fisher Scientific (2 mentions) Cd25 pe, by Thermo Fisher Scientific (1 mentions) Cd69 fitc, by Thermo Fisher Scientific (1 mentions) Anti mouse cd4 pe cy7, by Thermo Fisher Scientific (1 mentions) Fixable viability dye efluor 506, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

PerCP-Cy5.5 (62 citations) B220 PerCP-Cy5.5 (RA3-6B2, (3 citations) B220-PerCP (2 citations) PerCP-Cy5.5-anti-B220 (2 citations) CD45R-B220-PerCP-Cy5.5 (2 citations)

7 protocols using b220 percp cy5

1

Multiparameter Flow Cytometry Analysis

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Splenocytes, mesenteric lymph node cells, and inguinal lymph node cells were isolated from the indicated mouse and then stained with anti-mouse CD4-PE/Cy7, CD8-Percp/Cy5.5, CD25-PE, CD69-FITC, CD44-APC-Cy7, CD62L-FITC, B220-Percp/Cy5.5, GL-7-FITC, and CD95-PE antibodies (eBioscience, San Diego, CA, USA) for 15 min at 4 °C. For TFH cell analysis, cells were stained with anti-mouse CXCR5-biotin for 30 min at 4 °C followed by anti-mouse CD44-APC/Cy7 and CD4-PE/Cy7 and streptavidin-APC staining. After fixation and permeabilization using a Foxp3 Staining Kit (eBioscience), intracellular Bcl-6 was stained using anti-mouse Bcl-6-PE (BD Bioscience, San Jose, CA, USA) for 40 min at room temperature. Cells were examined using a FACSCanto II system (BD Bioscience), and data were analyzed using FlowJo software (Treestar, Ashland, OR, USA).
Kim D.H., Park H.J., Park H.S., Lee J.U., Ko C., Gye M.C, & Choi J.M. (2019). Estrogen receptor α in T cells suppresses follicular helper T cell responses and prevents autoimmunity. Experimental & Molecular Medicine, 51(4), 41.
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2

Multiparameter Analysis of B Cell Subsets

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Cells were incubated with Fc blockers (BioLegend) for 15 min, and then stained with the following antibodies for 45 min: B220-PerCp-Cy5.5, CD5-eF450, Cd11b-APC-eF780, IgM-APC, and Fixable Viability Dye eFluor® 506 (eBioscience) or Ghost Dye Violet 510 (Tonbo Biosciences) for live cell gating, CD5-AF647 (100614, BioLegend) and IgM-AF350 (Life Technologies) for microscopy. Cells were then washed three times by centrifugation. FACS data were collected with BD LSRII and analyzed using FlowJo software (TreeStar). Images were taken at 63X oil-immersion objective with Leica TCS SP5 microscope.
Vo H., Chiu J., Allaimo D., Mao C., Wang Y., Gong Y., Ow H., Porter T, & Zhong X. (2014). High fat diet deviates PtC-specific B1 B cell phagocytosis in obese mice. Immunity, Inflammation and Disease, 2(4), 254-261.
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3

Multi-Dimensional Flow Cytometry Profiling

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Splenocytes, mesenteric, and inguinal lymph node cells were isolated and then stained with anti-mouse CD4-APC, CD8-PerCP-Cy5.5, CD44-PE, CD62L-FITC, GL-7-FITC, CD95-PE, and B220-PerCP-Cy5.5 antibodies (eBioscience, San Diego, CA) for 15 min at 4 °C. For TFH differentiation analysis, the cells were stained with anti-mouse CXCR5-biotin for 30 min at 4 °C followed by anti-mouse CD44-FITC, CD4-PerCP-Cy5.5, and streptavidin-APC staining. After fixation and permeabilization using the Foxp3 Staining Kit (eBioscience), anti-mouse Bcl-6-PE was stained for 1 h at room temperature. Cells were examined using the FACSCanto II system (BD Bioscience, San Jose, CA, USA) and data were analyzed using Flow Jo software (Treestar, Ashland, OR, USA). In all cases, doublets (FSC-area versus FSC-height gating) were excluded.
Park H.J., Park H.S., Lee J.U., Bothwell A.L, & Choi J.M. (2016). Gender-specific differences in PPARγ regulation of follicular helper T cell responses with estrogen. Scientific Reports, 6, 28495.
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4

Adoptive Transfer of Mpzl3-Deficient Lymphocytes

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Bone marrow was flushed out of femurs and tibias of Mpzl3 −/− or +/+ littermates (6 weeks old). The bone marrow was labeled using biotinylated antibodies against CD3, CD4, CD8 and B220 (eBioscience, San Diego, CA). Streptavidin-labeled microbeads were then used to magnetically deplete the biotinylated lymphocytes using autoMACS (Milteyni Biotec, Inc., Auburn, CA). 3 × 106 bone marrow cells after depletion were injected intraperitoneally into 2-day old B6 Rag −/− mice (The Jackson Laboratory, Bar Harbor, ME). After 10 weeks, the inguinal lymph nodes and spleens of these mice were analyzed by flow cytometry, and skin analyzed by histology. For flow cytometry, 3 × 106 cells were blocked using a cocktail of anti-CD16/32 (2.4G2) and normal mouse sera (Jackson ImmunoResearch, West Grove, PA) before being stained with fluorescently labeled antibodies to determine their phenotype and activation status. The following antibody conjugates were used: CD11b PE-TR (Life Technologies Corp.), CD4 V500 (Becton Dickinson, San Jose, CA), CD11c FITC, Gr-1 PECy7, CD3 Alexa Fluor 700, CD8 efluor605, B220 PerCpCy5.5, CD44 efluor450, CD62L APC, CD69 PE (eBioscience). Cells were then washed and re-suspended in 2% FBS in PBS for analysis with flow cytometers (LSR-II and Fortessa, Becton Dickinson).
Leiva A.G., Chen A.L., Devarajan P., Chen Z., Damanpour S., Hall J.A., Bianco A.C., Li J., Badiavas E.V., Zaias J., Miteva M., Romanelli P., Nouri K, & Wikramanayake T.C. (2014). Loss of Mpzl3 Function Causes Various Skin Abnormalities and Greatly Reduced Adipose Depots. The Journal of investigative dermatology, 134(7), 1817-1827.
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5

Immunophenotyping and Cell Cycle Analysis

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Cells were stained using antibodies directed against: CD43-APC,
B220-FITC, CD19-APC-Cy7, CD43-PE, Streptavidin-APC-Cy7, IgM-FITC, CD16/CD32 Fc
receptor block (all from BD Biosciences), CD19-eFluor450, B220-PerCP-Cy5.5,
BP1-biotinylated and IgD-APC (all from eBiosciences). For cell-cycle analysis 1
x 106 cells were stained with propidium iodide (PI, 50 μg/ml)
in a hypotonic lysis buffer (0.1% sodium citrate, 0.1% triton X-100, 100
μg/ml RNAse) and incubated at 37°C for 30 minutes. Analysis of
apoptotic fractions was performed by staining with Annexin V and 7-AAD from the
Annexin V Apoptosis Detection Kit eFluor® 450 (eBiosciences)
following manufacturer’s instructions. Samples were analyzed using a FACS
Canto II flow cytometer equipped with 488, 633 and 405 nm lasers (BD
Biosciences). FACS-Plots and calculations were done using the FACS Diva software
version 6.1.2 (BD Biosciences). High-purity FACS-sorting was performed on a FACS
Aria III equipped with a 488 nm laser (BD Biosciences) as described (2 ).
Bellutti F., Tigan A.S., Nebenfuehr S., Dolezal M., Zojer M., Grausenburger R., Hartenberger S., Kollmann S., Doma E., Prchal-Murphy M., Uras I.Z., Höllein A., Neuberg D.S., Ebert B.L., Ringler A., Mueller A.C., Loizou J.I., Hinds P.W., Vogl C., Heller G., Kubicek S., Zuber J., Malumbres M., Farlik M., Villunger A., Kollmann K, & Sexl V. (2018). CDK6 antagonizes p53-induced responses during tumorigenesis. Cancer discovery, 8(7), 884-897.
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6

Multicolor Flow Cytometry Antibody Panel

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Commercial antibodies and staining reagents, from BioLegend: GL7-PacBlue, GL7-PE, GL7-A647, B220-PerCP-Cy5.5, B220-APC-Cy7, IgMb-FITC, IgMa-PE, CD45.2-FITC, CD45.2-APC, CD45.1-FITC, CD45.1-PE, IgD-PB, CD21/35 (7E9)-PE, CD138-PE, CD38-PE-Cy7, CD31-A647, CD157-PE, Streptavidin-PE/Cy7; from eBioscience: CD95 (APO-1/Fas)-PE (clone 15A7) and viability dye Fixable live/dead stain Efluor780; from ThermoFisher Scientific: Rabbit-anti-Goat-A488, Hoechst 33342, DAPI (4',6-Diamidino-2-Phenylindole, Dihydrochloride), Phalloidin-A568; from Southern Biotech: AP-Goat-anti-Mouse IgG, AP-Goat-anti-Mouse IgG2a, AP-Goat-anti-Mouse IgG2c; from Rockland Immunochemicals: Rabbit polyclonal anti-B-Phycoerythrin; from DAKO: Rabbit polyconal anti-Mouse Immunoglobulins-biotin; from Perkin-Elmer: Europium-labeled streptavidin. In-house generated anti-idiotypic Ab, clone 9D11, conjugated to Alexa Fluor 647 or Alexa Fluor 568; in-house generated Rabbit polyclonal anti-C3b conjugated to Alexa Fluor 633.
Degn S.E., van der Poel C.E., Firl D.J., Ayoglu B., Al Qureshah F.A., Bajic G., Mesin L., Reynaud C.A., Weill J.C., Utz P.J., Victora G.D, & Carroll M.C. (2017). Clonal Evolution of Autoreactive Germinal Centers. Cell, 170(5), 913-926.e19.
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7

Adoptive Transfer of Mpzl3-Deficient Lymphocytes

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Bone marrow was flushed out of femurs and tibias of Mpzl3 −/− or +/+ littermates (6 weeks old). The bone marrow was labeled using biotinylated antibodies against CD3, CD4, CD8 and B220 (eBioscience, San Diego, CA). Streptavidin-labeled microbeads were then used to magnetically deplete the biotinylated lymphocytes using autoMACS (Milteyni Biotec, Inc., Auburn, CA). 3 × 106 bone marrow cells after depletion were injected intraperitoneally into 2-day old B6 Rag −/− mice (The Jackson Laboratory, Bar Harbor, ME). After 10 weeks, the inguinal lymph nodes and spleens of these mice were analyzed by flow cytometry, and skin analyzed by histology. For flow cytometry, 3 × 106 cells were blocked using a cocktail of anti-CD16/32 (2.4G2) and normal mouse sera (Jackson ImmunoResearch, West Grove, PA) before being stained with fluorescently labeled antibodies to determine their phenotype and activation status. The following antibody conjugates were used: CD11b PE-TR (Life Technologies Corp.), CD4 V500 (Becton Dickinson, San Jose, CA), CD11c FITC, Gr-1 PECy7, CD3 Alexa Fluor 700, CD8 efluor605, B220 PerCpCy5.5, CD44 efluor450, CD62L APC, CD69 PE (eBioscience). Cells were then washed and re-suspended in 2% FBS in PBS for analysis with flow cytometers (LSR-II and Fortessa, Becton Dickinson).
Leiva A.G., Chen A.L., Devarajan P., Chen Z., Damanpour S., Hall J.A., Bianco A.C., Li J., Badiavas E.V., Zaias J., Miteva M., Romanelli P., Nouri K, & Wikramanayake T.C. (2014). Loss of Mpzl3 Function Causes Various Skin Abnormalities and Greatly Reduced Adipose Depots. The Journal of investigative dermatology, 134(7), 1817-1827.
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