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Ra3 6b2

Manufactured by Thermo Fisher Scientific
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Sourced in Germany, United States

The RA3-6B2 is a laboratory equipment designed for specific applications. It features a compact and durable construction. The core function of this product is to provide a reliable and versatile solution for laboratory operations. Further details on the intended use of this product are not available.

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Lab products found in correlation

Rb6 8c5, by Thermo Fisher Scientific (3 mentions) Apc cy7 anti cd45, by Thermo Fisher Scientific (2 mentions) Fitc pe pe cy7 anti ly6a e d, by Thermo Fisher Scientific (2 mentions) Bv421 pe pe cy7 anti cd117 2, by Thermo Fisher Scientific (2 mentions) Biotin anti cd48, by Thermo Fisher Scientific (2 mentions) Biotin anti cd41, by Thermo Fisher Scientific (2 mentions) Pe anti cd51, by Thermo Fisher Scientific (2 mentions) Alexa647 anti ve cadherin bv13 apc anti cd31, by Thermo Fisher Scientific (2 mentions) Fitc pe pe cy7 anti cd45, by Thermo Fisher Scientific (2 mentions) Pe anti cd150, by Thermo Fisher Scientific (2 mentions) Cd41 percp efluor 710, by Thermo Fisher Scientific (2 mentions) Brdu alexa 647 3d4, by BD (2 mentions) Alexa 647 anti ki67 sola 15, by Merck Group (2 mentions) Hoechst 33342, by Merck Group (2 mentions) Cd3 17a2, by Thermo Fisher Scientific (2 mentions) Prolong gold antifade mountant, by Thermo Fisher Scientific (2 mentions) Smz1270 stereo microscope, by Nikon (2 mentions) Superfrostplus adhesion glass, by Thermo Fisher Scientific (2 mentions) Inverted lsm780 microscope, by Zeiss (2 mentions) Plan apochromat 63 na 1.40 oil immersion objective, by Zeiss (2 mentions)

Close spelling variants of this product

B220 (clone RA3-6B2; (17 citations) FITC-labeled anti-B220 (clone RA3-6B2) (2 citations) B220-AF488 clone RA3–6B2 (2 citations) Anti-B220-APC (clone RA3-6B2, (3 citations) B220 PerCP-Cy5.5 (RA3-6B2, (3 citations) B220 (RA3–6B2) APC (4 citations) B220-FITC (RA3-6B2, (3 citations) Anti-B220/CD45R (clone RA3-6B2, (2 citations) α-CD45R (clone RA3-6B2 (2 citations) B220 (RA3–6B2)-APC/APC-Cy7 (2 citations)

30 protocols using ra3 6b2

1

Multicolor Flow Cytometry Panel for Immune Cell Profiling

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Cells were stained with the following fluorochrome-conjugated antibodies: anti-B220 (eBioscience, clone RA3-6B2), anti-CD3e (eBioscience, clone 145-2C11), anti-CD8a (BD, clone 53-6.7), anti-CD11b (eBioscience, clone M1/70), anti-CD11c (Bio Legend, clone N418), anti-CD14 (Bio Legend, clone SA14-2), anti-CD19 (eBioscience, clone eBio1D3), anti-CD64 (Bio Legend, clone X54-5/7.1), anti-CD68 (AbD Serotec, clone FA-11), anti-CD163 (Bioss, polyclonal), anti-CD115 (eBioscience, clone AF598), anti-CCR3 (Bio Legend, clone J073E5), anti-F4/80 (Bio Legend, clone CI:A3-1), anti-FPR-1 (Bioss, polyclonal), anti-MHC II (Bio Legend, clone M5/114.15.2), anti-MR (AbD Serotec, clone MR5D3) and anti-PILRa (R&D Systems, polyclonal). Fc receptors were blocked with 1.5mg/ml human IgG (Privigen). Dead cells were excluded using the Hoechst 33342 dye. Only events that appeared single in forward-scatter width were analyzed. The gating strategy is shown in Supplementary Figure 7. A FACSCanto II and FACSDiva software (BD) were used for flow cytometry and data were analyzed with FlowJo software (TreeStar).
Franken L., Klein M., Spasova M., Elsukova A., Wiedwald U., Welz M., Knolle P., Farle M., Limmer A, & Kurts C. (2015). Splenic red pulp macrophages are intrinsically superparamagnetic and contaminate magnetic cell isolates. Scientific Reports, 5, 12940.
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2

Multicolor Flow Cytometry Antibodies

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The following antibodies were used in this study: biotin-anti-Lineage (TER-119, RB6-8C5, RA3-6B2, M1/70, 145-2C11, 1:50 dilution), FITC/PE-anti-CD45.2 (104), FITC/PE/PE-Cy7-anti-Ly6A/E (D7), BV421/PE/PE-Cy7-anti-CD117 (2B8), biotin-anti-CD48 (HM48-1), biotin-anti-CD41 (MWReg30, 1:2500 dilution), APC-Cy7-anti-CD45 (30-F11), Pe-anti-CD51 (RMV-7), Alexa647-anti-VE-cadherin (BV13) APC-anti-CD31 (MEC13.3), FITC/PE/PE-Cy7-anti-CD45.1 (A20), PE-anti-CD150 (TC15-12F12.2), CD41-PerCP-eFluor 710 (MWReg30), Gr-1-FITC/Alexa 647 RB6-8C5), CD4-PE-Cy7 (GK1.5), CD8-PE-Cy7 (53-6.7), CD115-PE (AFS98), APC-eflour780 (RA3-6B2) all from eBioscience. CD16/32-APC-Cy7 (93), CD105-PE-Cy7 (MJ7/18), Ter119-Alexa 700 (Ter 119), F4/80-Alexa 647 (BM8,) were from Biolegend. BrdU-Alexa 647 (3D4, BD Biosciences, 1:50 dilution), Alexa 647-anti-Ki67 (SolA 15) and Hoechst 33342 (Sigma). Unless otherwise specified, all antibodies were used at a 1:100 dilution.
Bruns I., Lucas D., Pinho S., Ahmed J., Lambert M.P., Kunisaki Y., Scheiermann C., Schiff L., Poncz M., Bergman A, & Frenette P.S. (2014). Megakaryocytes regulate hematopoietic stem cell quiescence via Cxcl4 secretion. Nature medicine, 20(11), 1315-1320.
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3

Monoclonal Antibody Panel for Murine Immune Cell Characterization

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S12, a mouse anti-LMP1 immunoglobulin G (IgG) antibody (Ab) directed to the C-terminus of LMP1, was a gift from Dr. Fred Wang (Harvard University, Cambridge, MA). Monoclonal antibodies (mAbs) specific for rat IgG2a (eBR2a), mouse IgG2a κ (isotype control) (eBM2a), mouse/human CD45R (B220) (RA3-6B2), mouse CD21/CD35 (ebio4E3), mouse IgD (11–26), mouse CD11b (M1/70), mouse IgG2b κ control (eBMG2b), mouse CD5 (53-7.3), human/mouse terminal deoxynucleotidyl transferase TdT (19-3), mouse CD19 (MB19-1), mouse IgM (II/41), rat IgG1 control (eBRG1), mouse T- and B-cell activation antigen (GL-7), mouse CD16/32 (to block FcR binding), mouse CD23 (B3B4) and mouse C1qRp (AA4.1) were purchased from eBioscience (San Diego, CA). mAbs specific for mouse CD43 (S7), mouse Igλ1, λ2 and λ3 light chain (R26-46) and mouse Igκ light chain (187.1) were purchased from BD Biosciences (San Jose, CA). The anti-mouse CD40 mAb, 1C10, was produced from a hybridoma provided by Dr. Frances Lund (University of Alabama, Birmingham, AL). The isotype control mAb for 1C10, mAb72, was produced from the 72-2 hybridoma (rat IgG2a isotype control) obtained from the American Type Culture Collection (ATCC, Manassas, VA). Anti-mouse interleukin-6 (IL-6) mAbs (MP5-20F3 and MP5-32C11, biotin-conjugated) used for enzyme-linked immunosorbent assay (ELISA) were obtained from eBioscience.
Ontiveros E.P., Halwani A., Stunz L.L., Kamberos N., Olivier A.K., Janz S, & Bishop G.A. (2014). A new model of LMP1–MYC interaction in B cell lymphoma. Leukemia & lymphoma, 55(12), 2917-2923.
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4

Lymph Node Immunohistochemistry

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Cryostat sections (8 μm thick) of snap-frozen lymph nodes were air-dried, fixed in acetone for 10 min, and blocked with PBS containing 2% FCS. Sections were stained with the following anti-mouse antibodies in blocking solution for 45 min: monoclonal antibody to VSV glycoprotein (VSV-G, self-made), anti-CD169 (3D6.112, Acris Antibodies; 645608, R&D Systems), anti-CD45R (B220) (RA3-6B2, eBiosciences), and anti-F4/80 (BM8, eBiosciences). Imaging was performed on a KEYENCE BZ II analyzer fluorescence microscope.
Solmaz G., Puttur F., Francozo M., Lindenberg M., Guderian M., Swallow M., Duhan V., Khairnar V., Kalinke U., Ludewig B., Clausen B.E., Wagner H., Lang K.S, & Sparwasser T.D. (2019). TLR7 Controls VSV Replication in CD169+ SCS Macrophages and Associated Viral Neuroinvasion. Frontiers in Immunology, 10, 466.
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5

Comprehensive Phenotyping of Mouse Splenocytes

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2 × 106 freshly isolated mouse splenocytes, resuspended in PBS with 2% FBS, were labeled for 30 min at 4°C in the dark with different antibody panels. The antibodies used were: α-CD8 (5H10, Thermo Fisher, cat. nr. MCD0828TR), α-CD4 (RM4-5, Thermo Fisher, cat. nr. 56-0042-80), α-CD3 (17A2, eBioscience, cat. nr. 46-0032-80), α-CD62L (MEL-14, Biolegend, cat. nr. 104411), α-CD44 (IM7, Biolegend, cat. nr. 103005), α-PD-1 (29F-1A12, Biolegend, cat. nr. 135223), α-IL7R (A7R34, Biolegend, cat. nr. 135031), α-CTLA-4 (UC10-4B9, eBioscience, cat. nr. 12-1522-81), α-B220 (RA3-6B2, eBioscience, cat. nr. 47-0452-82), α-CD21 (eBio4E3, eBioscience, cat. nr. 48-0212-80), α-CD23 (B3B4, Biolegend, cat. nr. 101612), α-CD86 (GL1, eBioscience, cat. nr. 12-0862-81), α-CD69 (H1.2F3, eBioscience, cat. nr. 11-0691-81), α-IAb (AF6-120.1, eBioscience, cat. nr. 46-5320-80). The cells were then washed twice with PBS, resuspended in PBS with 2% FBS and data were acquired on an LSRII Flow Cytometer (BD Biosciences) and analyzed with FlowJo software, ver. 9.9.6 (Treestar, Inc).
Fenninger F., Han J., Stanwood S.R., Nohara L.L., Arora H., Choi K.B., Munro L., Pfeifer C.G., Shanina I., Horwitz M.S, & Jefferies W.A. (2019). Mutation of an L-Type Calcium Channel Gene Leads to T Lymphocyte Dysfunction. Frontiers in Immunology, 10, 2473.
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6

Multiparameter Flow Cytometry Assay

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Antibodies against murine CD4 (RM4-5, AF700, BV650), CD8 (53-6.7, APC/Fire 750, BV785), CCR2 (SA203G11, BV421), CD11c (N418, APCCy7, Pe/Cy5), CD206 (C068C2, APC), CD11b (M1/70, BV421), F4/80 (BM8, BV510, BV711, FITC), TCRβ (H57-597, BV605), and NK1.1 (PK136, BV650) were purchased from BioLegend. Antibodies against CD4 (GK1.5, BUV395) and ST2 (U29-93, BV480) were purchased from BD Biosciences. Antibodies against B220 (RA3-6B2, AF488), CD11c (N418, e450, PE-Cy5.5), CD25 (PC61.5, PE-Cy7), CD4 (GK1.5, APC-eFluor 780, Super Bright 645), CD45 (30-F11, Pacific Orange), Foxp3 (FJK-16s, APC, e450, FITC), GITR (DTA-1, Super Bright 600), IL-10 (JES5-16E3, PE), KLRG1 (2F1, APC, APC-eFluor 780, PE-eFluor 610), NK1.1 (PK136, PE), and ST2 (RMST2-2, PE, PerCP-eFluor710) were purchased from eBioscience, Thermo Fisher Scientific. Viability dyes (e506, near IR, UV) were purchased from Invitrogen, Thermo Fisher Scientific.
Extracellular flow staining was performed in 2% FBS-PBS in the presence of BioLegend’s TruStain FcX for 30 minutes at 4°C. Fixation, permeabilization, and intracellular staining were performed using eBioscience’s (Thermo Fisher Scientific) Foxp3/transcription factor staining buffer set per manufacturer’s directions. Flow data were collected using a Cytek Aurora or BD Biosciences LSR and analyzed using FlowJo.
Beppu L.Y., Mooli R.G., Qu X., Marrero G.J., Finley C.A., Fooks A.N., Mullen Z.P., Frias AB J.r., Sipula I., Xie B., Helfrich K.E., Watkins S.C., Poholek A.C., Ramakrishnan S.K., Jurczak M.J, & D’Cruz L.M. (2021). Tregs facilitate obesity and insulin resistance via a Blimp-1/IL-10 axis. JCI Insight, 6(3), e140644.
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7

Flow Cytometric Analysis of Immune Cell Subsets

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All cell experiments were strictly prepared on ice, unless otherwise stated in other specific procedures. Cells (1×106 cells/sample) were washed with fluorescence-activated cell sorting staining buffer (phosphate-buffered saline, 2% fetal bovine serum or 1% bovine serum albumin, 0.1% sodium azide). All samples were incubated with anti-Fc receptor Ab (clone 2.4G2, BD Biosciences, San Jose, CA), prior to incubation with other Abs diluted in fluorescence-activated cell sorting buffer supplemented with 2% anti-Fc receptor Ab. For intracellular cytokine staining, cells were collected and fixed for 50 min with 1mL fixation buffer (IC Fixation and Permeabilization kit, eBioscience, San Diego, CA). After washing, the fixed cells were stained. The samples were filtered immediately before analysis or cell sorting to remove any clumps. The following antibodies were used: fluorescence-conjugated anti-mouse B220 (eBioscience, RA3-6B2), CD3 (eBioscience, 145-2C11), CD4 (eBioscience, GK1.5), CD8 (Biolegend, 53-6.7), RORγt (eBioscience, B2D), IL-17A (eBioscience, eBio17B7), CD24 (eBioscience, M1/69), CD138 (Biolegend, 281-2), and IgG (Santa Cruz Biotech, F1306) antibodies. Data collection and analyses were performed on a FACS Calibur flow cytometer using CellQuest software.
Xing C., Zhu G., Xiao H., Fang Y., Liu X., Han G., Chen G., Hou C., Shen B., Li Y., Ma N, & Wang R. (2017). B cells regulate thymic CD8+T cell differentiation in lupus-prone mice. Oncotarget, 8(52), 89486-89499.
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8

Multimodal Imaging of Immune Cells

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For immunofluorescence, spleens were embedded and frozen in OCT, sectioned at 15 μm and mounted on SuperFrostPlus Adhesion glass (Thermo Fisher Scientific). Sections were dehydrated using silica beads, fixed with 4% paraformaldehyde for 10 min and washed with PBS. Samples were blocked using 5% normal goat serum for 2 h before staining. Samples were incubated with antibodies against B220 (RA3-6B2, eBioscience), CD3 (17A2, eBioscience) and F4/80 (BM8, Biolegend) diluted in 5% NGS for 2 h at room temperature in the dark. After staining, samples were washed with PBS at least three times. Samples were then mounted using ProLong Gold Antifade Mountant (Invitrogen) and imaged using an inverted LSM780 microscope (Carl Zeiss) and a plan apochromat 63× NA 1.40 oil-immersion objective (Carl Zeiss). For haematoxylin and eosin (H&E) staining, organs were collected and fixed in 10% formalin. Fixed samples were embedded in paraffin and sectioned at 10 μm, mounted on SuperFrostPlus Adhesion glass and stained using H&E. Mounted samples were imaged using a Nikon SMZ1270 Stereo Microscope. Imaging data were analysed using Fiji (ImageJ) software (NIH).
Tsui C., Kretschmer L., Rapelius S., Gabriel S.S., Chisanga D., Knöpper K., Utzschneider D.T., Nüssing S., Liao Y., Mason T., Torres S.V., Wilcox S.A., Kanev K., Jarosch S., Leube J., Nutt S.L., Zehn D., Parish I.A., Kastenmüller W., Shi W., Buchholz V.R, & Kallies A. (2022). MYB orchestrates T cell exhaustion and response to checkpoint inhibition. Nature, 609(7926), 354-360.
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9

Multiparametric Immune Cell Profiling

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Blood cells were collected from the eye vein of anesthetized mice, and splenocytes from the spleen after filtration through cell strainers (70 µm). Ammonium-chloride-potassium lysing buffer was used for the lysis of red blood cells. Blood cells were labeled for 15 min at 4°C with fluorochrome-conjugated monoclonal antibodies: CD45-FITC BD Biosciences Clone 30-F11 (1:100), Ly6G BV421 BioLegend Clone 1A8 (1:500), Ly6C BV605 BD Biosciences Clone AL-21 (1:500), CD3e BV786 BD Biosciences Clone 5OOA2 (1:500), CD45R (B220) PE BD Biosciences Clone RA3-6B2 (1:500) and CD11b PE-Cy7 eBioscience Clone M1/70 (1:500). Splenocytes were labeled for 15 min at 4°C with fluorochrome-conjugated monoclonal antibodies: CD3 BV786 BD Biosciences (1:500), CD11b BV510 BD Biosciences (1:500), CD49b PE eBioscience (1:500), CD4 BV650 BD Biosciences (1:500), CD25 FITC BioLegend (1:500) and Foxp3 PerCP-Cy5.5 eBioscience (1:100). Samples were acquired with a BD LSRII Fortessa (BD Biosciences) and analyzed with FlowJo software.
Cordero-Sanchez C., Riva B., Reano S., Clemente N., Zaggia I., Ruffinatti F.A., Potenzieri A., Pirali T., Raffa S., Sangaletti S., Colombo M.P., Bertoni A., Garibaldi M., Filigheddu N, & Genazzani A.A. (2019). A luminal EF-hand mutation in STIM1 in mice causes the clinical hallmarks of tubular aggregate myopathy. Disease Models & Mechanisms, 13(2), dmm041111.
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10

Western Blot and Flow Cytometry Analysis

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The following Abs were used for Western blot analyses or flow cytometry analysis: p53 (DO-1, Santa Cruz), p21 (DSC60, Cell Signaling Technology, Danvers, MA), β-actin (AC-15, Sigma-Aldrich, St. Louis), His (Invitrogen), GST (B-14, Santa Cruz), HA (F-7, Santa Cruz), HA (C29F4, Cell Signaling Technology), Myc (9E10, Santa Cruz), Ab against mouse DD1α (MH5A, BioLegend), Ab against human PD-1 (J116, eBioscience), Ab against human PD-L1 (M1H1, eBioscience), CTLA4 (C-19, Santa Cruz), ICOSL (LifeSpan BioSciences), Ab against human CD14 APC (61D3, eBioscience), Ab against mouse F4/80 APC (BM8, eBioscience), Ab against mouse CD45R/B220 (RA3–6B2, eBioscience), Ab against human CD4-APC (OKT4, eBioscience), Ab against mouse CD4-APC (RM4–5, eBioscience), Ab against human CD8-APC (SK1, eBioscience) and Ab against mouse CD8a+-APC (53–6.7, eBioscience). Rabbit polyclonal and mouse monoclonal Ab against DD1α was raised against human DD1α (amino acids 33–311) as an immunogen. Beads immobilized with Abs against HA (Roche) or Myc (9B11, Cell Signaling) were used for immunoprecipitations. Recombinant human PD-1–Ig, mouse PD-1–Ig, human PD-L1–Ig, mouse PD-L1–Ig, and control Ig proteins were from R&D Systems.
Yoon K.W., Byun S., Kwon E., Hwang S.Y., Chu K., Hiraki M., Jo S.H., Weins A., Hakroush S., Cebulla A., Sykes D.B., Greka A., Mundel P., Fisher D.E., Mandinova A, & Lee S.W. (2015). Control of signaling-mediated clearance of apoptotic cells by the tumor suppressor p53. Science (New York, N.Y.), 349(6247), 1261669.
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