CD141 (BDCA-3) Antibody, anti-human, REAfinity™
Clone:
REA674
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
FC, MICS, IF, IHC, MC
Alternative names:
Thrombomodulin, Fetomodulin, TM

Extended validation for CD141 (BDCA-3) Antibody, anti-human, REAfinity™

Specificity

Epitope competition
In order to compare the epitope specificity of an antibody, the clone being used is compared with other known clones recognizing the same antigen in a competition assay.
Other clonesOverlap in epitope recognition with REA674
501733-
1A4-
AD5-14H12++
JAA17-
M80-
Cells were incubated with an excess of purified unconjugated CD141 (BDCA-3) (REA674) antibody followed by staining with fluorochrome-conjugated antibodies of other known clones against the same marker. Based on the fluorescence signal obtained, the clones were identified as recognizing completely overlapping (++), partially overlapping (+), or completely different epitopes (-) of the marker.
Knockout validation
To ensure antibody specificity, the target gene is knocked out in a suitable cell line using the CRISPR/Cas9 system and the knockout is confirmed by sequencing of the target locus. The antibody is considered to bind specifically to the intended epitope if no antibody binding to the knockout cells can be detected. The antibody staining is controlled by fluorescence microscopy and/or flow cytometry.
WT
KO
View details
Fluorescence microscopy image of CD141 (BDCA-3) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD141 (BDCA-3)-PE (REA674, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Fluorescence microscopy image of CD141 (BDCA-3) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD141 (BDCA-3)-PE (REA674, red) and counterstained with DRAQ5 (blue) as DNA stain.
Fluorescence microscopy image of CD141 (BDCA-3) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD141 (BDCA-3)-PE (REA674, red) and counterstained with DRAQ5 (blue) as DNA stain.

Sensitivity

Performance comparison
Selected fluorochrome conjugated antibodies from Miltenyi Biotec were compared to commercially available hybridoma clones in flow cytometry analysis.
View details
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
Flow cytometric comparison of different clones for CD141 (BDCA-3). Human peripheral blood mononuclear cells (PBMCs) were stained with CD141 (BDCA-3) antibodies and with a suitable counterstaining. As a control, CD141 (BDCA-3) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
Fixation data
To provide an indication on how an antibody performs after fixation of cells, in-house data on staining results before and after fixation with 3.7% formaldehyde using Miltenyi Biotec antibodies are provided. Different experimental settings may lead to different results.
No fixation
Post fixation
View details
Comparison of staining pattern on non-fixed and fixed cells using CD141 (BDCA-3) (REA674). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.
View details
Comparison of staining pattern on non-fixed and fixed cells using CD141 (BDCA-3) (REA674). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.
Comparison of staining pattern on non-fixed and fixed cells using CD141 (BDCA-3) (REA674). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.

Specifications for CD141 (BDCA-3) Antibody, anti-human, REAfinity™

Overview

Clone REA674 recognizes the human CD141 (BDCA-3) antigen, which is expressed at high levels on a minor subpopulation of human myeloid dendritic cells (about 0.02% of blood leukocytes). CD141 (BDCA-3)
high
blood dendritic cells are CD11c
dim
, CD123
–
, CD4
+
, Lin
–
, CD45RO
+
, CD2
–
, and CD16
–
. They express myeloid lineage markers, such as CD13 and CD33, and have a monocytoid morphology. Unlike CD1c (BDCA-1)
+
blood dendritic cells, CD141 (BDCA-3)
high
blood dendritic cells lack expression of CD2 and Fc receptors such as CD32, CD64, or FcεRI. CD141 (BDCA-3) is also present at very low levels on CD14
+
monocytes, granulocytes, CD303 (BDCA‑2)
+
CD304 (BDCA-4/Neuropilin-1)
+
plasmacytoid, and CD1c (BDCA-1)
+
myeloid dendritic cells. CD141 (BDCA-3)
high
CD1c (BDCA-1)
–
myeloid dendritic cells have been designated type-2 myeloid dendritic cells (MDC2s). CD141 is also known as thrombomodulin; thrombomodulin mediates co-agglutination by interaction with thrombin and protein C, though nothing is known about its function on MDC2s.
Additional information: Clone REA674 displays negligible binding to Fc receptors.

Alternative names

Thrombomodulin, Fetomodulin, TM

Detailed product information

Technical specifications

CloneREA674
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody (S), human IgG1
Hosthuman cell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Specieshuman
AntigenCD141 (BDCA-3)
Alternative names of antigenThrombomodulin, Fetomodulin, TM
Molecular mass of antigen [kDa]59
Distribution of antigendendritic cells, granulocytes, monocytes
Entrez Gene ID7056
RRIDAB_2751167, AB_2751233, AB_2751168, AB_2751230, AB_2751165, AB_2751235, AB_2751170, AB_2751234, AB_2751169, AB_2857380, AB_2857379, AB_2783957, AB_2783956, AB_2751236, AB_2751171, AB_2751231, AB_2751166, AB_2811443, AB_2811437, AB_2811645, AB_2751232

Resources for CD141 (BDCA-3) Antibody, anti-human, REAfinity™

Certificates

Please follow this
link
to search for Certificates of Analysis (CoA) by lot number.

References for CD141 (BDCA-3) Antibody, anti-human, REAfinity™

Publications

  1. Gosh, M. et al. (2012) CD13 regulates dendritic cell cross-presentation and T cell responses by inhibiting receptor-mediated antigen uptake. J Immunol 188(11): 5489-5499
  2. Mittag, D. et al. (2011) Human dendritic cell subsets from spleen and blood are similar in phenotype and function but modified by donor health status. J Immunol 186(11): 6207-6217
  3. Dzionek, A. et al. (2000) BDCA-2, BDCA-3, BDCA-4: Three markers for distinct subsets of dendritic cells in human peripheral blood. J Immunol 165: 6037-6046
  4. Chehimi, J. et al. (2002) Persistent decreases in blood plasmacytoid dendritic cell number and function despite effective highly active antiretroviral therapy and increased blood myeloid dendritic cells in HIV-infected individuals. J Immunol 168: 4769-4801
  5. Bosco, M. C. et al. (2011)
    Hypoxia modulates the gene expression profile of immunoregulatory receptors in human mature dendritic cells: identification of TREM-1 as a novel hypoxic marker
    in vitro
    and
    in vivo
    .
    Blood 117(9): 2625-2639
  6. Dutertre, C. A. et al. (2012)
    Pivotal role of M-DC8
    +
    monocytes from viremic HIV-infected patients in TNFα overproduction in response to microbial products.
    Blood 120(11): 2259-2268
  7. van de Ven, R. et al. (2011) Characterization of four conventional dendritic cell subsets in human skin-draining lymph nodes in relation to T-cell activation. Blood 118(9): 2502-2510
  8. MacDonald, K. P. A. et al. (2002) Characterization of human blood dendritic cell subsets. Blood 100: 4512-4520
  9. Chu, C. C. et al. (2012)
    Resident CD141 (BDCA3)
    +
    dendritic cells in human skin produce IL-10 and induce regulatory T cells that suppress skin inflammation.
    J. Exp. Med. 209(5): 935-945
  10. Jongbloed, S. L. et al. (2010)
    Human CD141
    +
    (BDCA-3)
    +
    dendritic cells (DCs) represent a unique myeloid DC subset that cross-presents necrotic cell antigens.
    J. Exp. Med. 207(6): 1247-1260
  11. Hémont, C. et al. (2013) Human blood mDC subsets exhibit distinct TLR repertoire and responsiveness. J. Leukoc. Biol. 93(4): 599-609
  12. Vanders, R. L. et al. (2013) Plasmacytoid dendritic cells and CD8 T cells from pregnant women show altered phenotype and function following H1N1/09 infection. J. Infect. Dis. 208(7): 1062-1070
  13. Jardine, L. et al. (2013) Rapid detection of dendritic cell and monocyte disorders using CD4 as a lineage marker of the human peripheral blood antigen-presenting cell compartment. Front Immunol 4: 495

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