Clone:
REA730
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
FC, MC
Alternative names:
PECAM-1, EndoCAM, GPIIa, PECA1

Extended validation for CD31 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 REA730
MBC.78:2-
L133.1-
WM59++
MEM-05-
1F11-
M89D3-
Cells were incubated with an excess of purified unconjugated CD31 (REA730) 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 CD31 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD31-PE (REA730, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Fluorescence microscopy image of CD31 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD31-PE (REA730, red) and counterstained with DRAQ5 (blue) as DNA stain.
Fluorescence microscopy image of CD31 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD31-PE (REA730, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Overlay histogram showing flow cytometric analysis of human CD31 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD31-PE, clone (REA730). 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 propidium iodide fluorescence.
Overlay histogram showing flow cytometric analysis of human CD31 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD31-PE, clone (REA730). 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 propidium iodide fluorescence.

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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31. Human peripheral blood mononuclear cells (PBMCs) were stained with CD31antibodies and with a suitable counterstaining. As a control, CD31 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 CD31 (REA730). 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 CD31 (REA730). 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 CD31 (REA730). 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 CD31 Antibody, anti-human, REAfinity™

Overview

Clone REA730 recognizes the human CD31 antigen, also known as PECAM-1. CD31 is a member of the immunoglobulin superfamily and is involved in leukocyte transmigration, angiogenesis, and integrin activation. CD31 is a 120–140 kDa single-chain transmembrane glycoprotein expressed on endothelial cells, monocytes, platelets, granulocytes, and several lymphocyte subsets. During maturation of CD4
+
T cells, the expression of CD31 changes: CD4
+
recent thymic emigrants (RTEs) express CD31 in contrast to central naive T cells. CD31 is highly expressed on endothelial cells, especially at their juncture.
Additional information: Clone REA730 displays negligible binding to Fc receptors.

Alternative names

PECAM-1, EndoCAM, GPIIa, PECA1

Detailed product information

Technical specifications

CloneREA730
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody (S), human IgG1
Hostcell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Specieshuman
AntigenCD31
Alternative names of antigenPECAM-1, EndoCAM, GPIIa, PECA1
Molecular mass of antigen [kDa]80
Distribution of antigenplatelets, B cells, NK cells, T cells, monocytes, granulocytes, endothelial cells
Entrez Gene ID5175
RRIDAB_2657279, AB_2657280, AB_2657281, AB_2657282, AB_2657283, AB_2657284, AB_2657285, AB_2657286, AB_2657287, AB_2657288, AB_2657289, AB_2657290, AB_2657291, AB_2657292, AB_2657293, AB_2657294, AB_2657295, AB_2819612, AB_2657278

Resources for CD31 Antibody, anti-human, REAfinity™

Certificates

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

References for CD31 Antibody, anti-human, REAfinity™

Publications

  1. Newman, P. J. et al. (1990) PECAM-1 (CD31) cloning and relation to adhesion molecules of the immunoglobulin gene superfamily. Science 247(4947): 1219-1222
  2. Stockinger, H. et al. (1990) Molecular characterization and functional analysis of the leukocyte surface protein CD31. J. Immunol. 145(11): 3889-3897
  3. Kimmig, S. et al. (2002) Two subsets of naive T helper cells with distinct T cell receptor excision circle content in human adult peripheral blood. J. Exp. Med. 195: 789-794

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