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Anti igm pe

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Anti-IgM-PE is a fluorochrome-conjugated antibody that binds to the Fc region of human immunoglobulin M (IgM) molecules. It is commonly used in flow cytometry and other immunoassays to detect and quantify IgM-positive cells or IgM levels in biological samples.

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Lab products found in correlation

Anti b220 percp, by BD (4 mentions) Anti b220 fitc, by BD (3 mentions) Anti cd25 pe, by BD (3 mentions) Anti cd5 pe, by BD (3 mentions) Facscalibur, by BD (3 mentions) Pe anti cd138, by BD (3 mentions) Anti cd43 pe, by BD (3 mentions) Cellquest, by BD (2 mentions) Anti cd4 fitc, by BD (2 mentions) Anti cd69 pe, by BD (2 mentions) Pe anti igκ, by BD (2 mentions) Anti cd86 pe, by BD (2 mentions) Anti cd80 pe, by BD (2 mentions) Anti igκ, by Southern Biotech (2 mentions) Anti mouse ceacam1 mab, by BD (2 mentions) Pe anti h 2kd, by BD (2 mentions) Pe anti 1 ad, by BD (2 mentions) Fluo 4, by Dojindo Laboratories (2 mentions) A20 cell line, by American Type Culture Collection (2 mentions) Pe anti cd3ε, by BD (2 mentions)

Spelling variants of this product

IgM-PE (8 citations) PE labeled anti-mouse IgM (2 citations) Rat anti-mouse anti-IgM-PE-Cy7 (2 citations) Anti-IgM-PE-Cy7 (2 citations) PE-Cy7-conjugated rat anti-mouse IgM (2 citations) PE-conjugated anti IgM (2 citations)

8 protocols using anti igm pe

1

Immunophenotypic Isolation of Murine B Cell Subsets

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Bone marrow was first depleted of red blood cells and B cells were
then purified with anti-B220 magnetic beads using the IMag system (BD
Pharmingen). B cells were subsequently stained with anti-IgM-PE,
anti-B220-PerCP or anti-B220-FITC, and anti-CD93/AA4.1-biotin or
anti-CD93/AA4.1-APC (BD Biosciences or Tonbo Biosciences). The biotinylated
antibody was detected with streptavidin-APC (Caltag or BD Biosciences).
Immature (B220+, IgM+, AA4.1+) and pre (B220+, IgM-, AA4.1+) B cells were
sorted on a FACS Aria (Becton Dickinson) or a MoFlo (Cytomation) cell
sorter. Samples were kept at 4° at all times prior to and during the
sort and until stimulation to avoid activating the BCR with the sorting
antibodies (11 (link)). Purified cells were
either harvested immediately or stimulated with 10 μg/ml goat
anti-mouse IgM F(ab’)2 (Jackson ImmunoResearch
Laboratories) for the indicated times at 106 cells/ml in RPMI +
10% FBS. Alternatively, bone marrow B lineage cells were expanded by
culturing for 4–6 days in 10 ng/ml IL-7 (R & D Systems) at 2
× 106 cells/ml and pre and immature B cells sorted as
above.
Ottens K., Hinman R.M., Barrios E., Skaug B., Davis L.S., Li Q.Z., Castrillon D.H, & Satterthwaite A.B. (2018). Foxo3 promotes apoptosis of BCR-stimulated immature B cells, thus limiting the window for receptor editing. Journal of immunology (Baltimore, Md. : 1950), 201(3), 940-949.
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2

Flow Cytometry Analysis of Immune Cells

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After the cells had been prepared, between 500,000 and 1.25 million cells from these single-cell suspensions were incubated with 1 μg of the appropriate primary antibodies (listed below) for 45 minutes on ice, with vortexing at every 7–10 minute intervals. These cells were washed twice with phosphate buffer followed by incubation for 15 minutes with an appropriate second step reagent-conjugated to a fluorochrome. After 15 minutes the cells were washed twice with PBS, and re-suspended in 1XPBS and fixed with paraformaldehyde at 1% final concentration. 10,000 and 25,000 cells were analyzed with a Becton Dickinson FACS Calibur (BD Biosciences, San Jose, CA, USA). Primary antibodies used in this investigation were: Anti-CD3-FITC, anti-CD3-PE anti-CD4-FITC, anti-CD4-PE, anti-CD44 Biotin, anti-B220-FITC and anti-IgM-PE (BioLegend Inc., San Diego, CA, USA). Also anti-CD25-PE, anti-CD5-PE, anti-IgM-PE, anti-IgA-FITC, and anti-B220-PerCP (BD Biosciences, San Jose, CA, USA). Secondary antibodies used were: Streptavidin-PErCP (BioLegend Inc, San Diego, CA, USA) and Streptavidin-PE (BD Biosciences, San Jose, CA, USA).
Jones M.A., DeWolf S., Vacharathit V., Yim M., Spencer S, & Bamezai A.K. (2016). Investigating B Cell Development, Natural and Primary Antibody Responses in Ly-6A/Sca-1 Deficient Mice. PLoS ONE, 11(6), e0157271.
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3

Flow Cytometric Immunophenotyping of Murine Immune Cells

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Bone marrow cells, splenocytes, and peritoneal wash cells were depleted of red blood cells and stained with various combinations of the following antibodies. Bone marrow: anti-CD43 FITC (BD Biosciences), anti-IgM PE (BD Biosciences), anti-B220 PerCP-Cy5.5 (Tonbo Biosciences), and anti-CD93 APC (Invitrogen). Spleen: anti-CD21 FITC (BD Biosciences), anti-CD23 PE (BD Biosciences), anti-CD95 PE (BD Biosciences), anti-IgM PErCP-Cy5.5 (BD Biosciences), B220 PerCP-Cy5.5, anti-B220 APC (Tonbo Biosciences), or anti-GL7 APC (BD Biosciences). Peritoneal wash: anti-CD11b FITC (BD Biosciences), anti-CD5 PE (Tonbo Biosciences), anti-IgM PErCP-Cy5.5, anti-B220 APC. Cultured B cells were stained with anti-CD138 PE (BD Biosciences), anti-B220 PerCP-Cy5.5, and anti-IgG1 biotin (BD Biosciences) plus streptavidin APC (Tonbo Biosciences). Samples were run on a FACS Calibur (BD) and analyzed with Flowjo (Treestar).
Ottens K., Schneider J., Kane L.P, & Satterthwaite A.B. (2020). PIK3IP1 promotes extrafollicular class switching in T-dependent immune responses. Journal of immunology (Baltimore, Md. : 1950), 205(8), 2100-2108.
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4

Assaying B cell development and proliferation

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For class switching assays, cell suspensions were stained with fluorochrome-conjugated anti-IgG1, anti-IgG3 (BD-Biosciences), anti-IgG2b (BioLegend), or anti-IgA (Southern Biotech). Samples were acquired on a LSRFortessa cell analyzer (BD-Biosciences). Analysis of B cell development and differentiation was performed using anti-CD21/CD35-FITC, anti-IgD-FITC, anti-IgM-PE, anti-IgM-FITC, anti-CD43-PE (BD-Biosciences) and anti-CD23-PE, anti-CD3-PE and anti-CD19-APC (BioLegend) antibodies. For cell proliferation analysis by cell tracking dye dilution, primary B cells were pulsed with 2 μM carboxyfluorescein succinimidyl ester (CFSE) (Invitrogen) or 5 μM CellTrace Violet (Thermofisher) for 10 min at 37C. CFSE/CellTrace covalently labels intracellular molecules, and each cell division halves the signal intensity. For cell cycle analysis, CH12 cells were collected, fixed, and permeabilized using Fixation/Permeabilization Solution (included in BrdU Flow Kit, BD-Biosciences) according to the manufacturer’s instructions. BrdU pulse and staining was performed by using BrdU Flow Kit (BD-Biosciences) according to the manufacturer’s instructions.
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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5

CLL Cell Apoptosis Analysis by Flow Cytometry

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Following treatment, CLL cells were harvested, stained with FITC/APC-conjugated Annexin V and 7-amino-actinomycin D (7-AAD). Flow cytometry data were acquired using a FACSCantoII flow cytometer (BD Biosciences) using the FACS Diva software package and analysed using the FlowJo software package (Tree Star, Inc., Ashland, OR) (22 (link)). Annexin V−7-AAD− cells were considered viable. For CLL-like mouse cells, anti-IgM-PE, anti-CD45-PerCP, anti-CD23-PE-Cy7, anti-CD5-APC, anti-CD19-APC-Cy7 and anti-CD11b-Pacific Blue abs were used (BD Biosciences). For intracellular staining, surface anti-B220-PE ab staining was followed by fixation with cytofix/cytoperm (BD Biosciences). Cells were then stained with anti-pAKT S473-AF647 and pS6S235/236-V450 abs.
Cosimo E., Tarafdar A., Moles M.W., Holroyd A.K., Malik N., Catherwood M.A., Hay J., Dunn K.M., Macdonald A.M., Guichard S.M., O’Rourke D., Leach M.T., Sansom O.J., Cosulich S.C., McCaig A.M, & Michie A.M. (2018). AKT/mTORC2 inhibition activates FOXO1 function in CLL cells reducing B cell receptor-mediated survival. Clinical cancer research : an official journal of the American Association for Cancer Research, 25(5), 1574-1587.
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6

Flow Cytometry Analysis of PBMCs

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All FACS measurements were conducted using a FACS Celesta (BD Biosciences, Franklin Lakes, NJ, USA). PBMC were thawed rapidly at 37°C. After washing with cold PBS, cells were counted using a Neubauer counting chamber and diluted to desired concentrations. For FACS analysis, PBMC were blocked with human serum for 30 min at 4°C and incubated with a pre-titrated antibody cocktail including RTX-AF647 as well as AF750 (Thermo Fisher Scientific), for live-dead staining. For FACS analysis, in each sample at least 200,000 cells were regularly acquired. If not otherwise indicated, all antibodies were obtained from BioLegend (San Diego, CA, USA). The following antibodies and fluorophores were used: V450 anti-CD27 (clone: M-T271), V500 anti-IgD (clone: IA6-2), BV650 anti-CD3 (clone: OKT3), BV785 anti-CD45 (clone: HI30), FITC anti-CD38 (clone: HIT2; BD BioSciences), PE anti-IgM (clone: MHM-88), PerCP anti-CD4 (clone: L200), PE-Cy7 anti-CD19 (clone: HIB19), AF700 anti-CD8a (clone: HIT8a). PBMC were washed twice and measured using a FACS Celesta (BD BioSciences). Graphical analysis was performed using FlowJo version 10.6.1 (FlowJo, Ashland, OR, USA). Of note, in contour plots not all cells are depicted as single dots. For gating strategy see supplemental information (Figure S1).
Webendörfer M., Reinhard L., Stahl R.A., Wiech T., Mittrücker H.W., Harendza S, & Hoxha E. (2021). Rituximab Induces Complete Remission of Proteinuria in a Patient With Minimal Change Disease and No Detectable B Cells. Frontiers in Immunology, 11, 586012.
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7

Characterization of A20 B cell lymphoma

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A20 cell line, derived from mouse B cell lymphoma on BALB/c background [13 (link)], was obtained from American Type Culture Collection (Manassas, VA). Primary cells were isolated from bone marrow (BM), spleen (SPL) or peripheral blood (PB) of B6 by standard methods. The A20 transfectants or primary cells were stained with the following antibodies (Abs): anti-mouse CEACAM1 mAb, CC1 (kindly provided by Dr. Kathlyn Holmes, University of Colorado, CO), anti-IgG1, FITC-anti-B220, PE-anti-IgM, FITC-anti-CD4, PE-anti-CD3ε, PerCP-anti-B220, PE-anti-CD43, PE-anti-CD25, PE-anti-Igκ, PE-anti-CD138, PE-anti-CD5, PE-anti-H-2Kd, PE-anti-I-Ad, PE-anti-CD69, PE-anti-CD80 and PE-anti-CD86 Abs (BD Biosciences, San Jose, CA). Data acquisition was performed using FACS Calibur™ and analysis software Cell Quest™ (BD Biosciences).
In some experiments, the A20 transfectants were treated with Fluo-4® (Dojindo, Kumamoto, Japan), and applied for FACS to analyze intracellular Ca2+ influx. During the acquisition, cells were treated with anti-Igκ (Southern Biotech, Birmingham, AL) to stimulate BCR signaling, and kinetics for Ca2+ influx were measured at FL2.
Tsugawa N., Yamada D., Watabe T., Onizawa M., Wang S., Nemoto Y., Oshima S., Tsubata T., Adachi T., Kawano Y., Watanabe M., Blumberg R.S., Okamoto R, & Nagaishi T. (2020). CEACAM1 specifically suppresses B cell receptor signaling-mediated activation. Biochemical and biophysical research communications, 535, 99-105.
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8

Characterization of A20 B cell lymphoma

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A20 cell line, derived from mouse B cell lymphoma on BALB/c background [13 (link)], was obtained from American Type Culture Collection (Manassas, VA). Primary cells were isolated from bone marrow (BM), spleen (SPL) or peripheral blood (PB) of B6 by standard methods. The A20 transfectants or primary cells were stained with the following antibodies (Abs): anti-mouse CEACAM1 mAb, CC1 (kindly provided by Dr. Kathlyn Holmes, University of Colorado, CO), anti-IgG1, FITC-anti-B220, PE-anti-IgM, FITC-anti-CD4, PE-anti-CD3ε, PerCP-anti-B220, PE-anti-CD43, PE-anti-CD25, PE-anti-Igκ, PE-anti-CD138, PE-anti-CD5, PE-anti-H-2Kd, PE-anti-I-Ad, PE-anti-CD69, PE-anti-CD80 and PE-anti-CD86 Abs (BD Biosciences, San Jose, CA). Data acquisition was performed using FACS Calibur™ and analysis software Cell Quest™ (BD Biosciences).
In some experiments, the A20 transfectants were treated with Fluo-4® (Dojindo, Kumamoto, Japan), and applied for FACS to analyze intracellular Ca2+ influx. During the acquisition, cells were treated with anti-Igκ (Southern Biotech, Birmingham, AL) to stimulate BCR signaling, and kinetics for Ca2+ influx were measured at FL2.
Tsugawa N., Yamada D., Watabe T., Onizawa M., Wang S., Nemoto Y., Oshima S., Tsubata T., Adachi T., Kawano Y., Watanabe M., Blumberg R.S., Okamoto R, & Nagaishi T. (2020). CEACAM1 specifically suppresses B cell receptor signaling-mediated activation. Biochemical and biophysical research communications, 535, 99-105.
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