Clone:
REA112
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
MC, FC, MICS, IF, IHC
Alternative names:
2B4, NAIL, NKR2B4, Nmrk, SLAMF4

Extended validation for CD244 (2B4) 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 REA112
C1.7++
2-69-
DM244++
Cells were incubated with an excess of purified unconjugated CD244 (2B4) (REA112) 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 CD244 (2B4) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD244 (2B4)-PE (REA112, red) and counterstained with DRAQ5 (blue) as DNA stain.
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Fluorescence microscopy image of CD244 (2B4) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD244 (2B4)-PE (REA112, red) and counterstained with DRAQ5 (blue) as DNA stain.
Fluorescence microscopy image of CD244 (2B4) knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD244 (2B4)-PE (REA112, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Overlay histogram showing flow cytometric analysis of CD244 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD244-PE, clone (REA112). 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 CD244 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD244-PE, clone (REA112). 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.
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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 CD244 (2B4) (REA112). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.
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Comparison of staining pattern on non-fixed and fixed cells using CD244 (2B4) (REA112). 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 CD244 (2B4) (REA112). 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 CD244 (2B4) Antibody, anti-human, REAfinity™

Overview

Clone REA112 recognizes CD244, a type I transmembrane receptor belonging to the CD2 subfamily of the Ig superfamily. Expression of CD244 is found on all natural killer (NK) cells, large subset of CD8
+
T cells, a small fraction of CD4
+
T cells, and eosinophils. Human CD244 upon ligand engagement activates NK cell cytotoxicity and production of interferon gamma (IFN-γ). Activating function of human CD244 requires recruitment of SH2 domain containing molecule called SLAM associated protein (SAP). In humans, CD244 has also been implicated as an inhibitory receptor such as in developing NK cells. In contrast, murine CD244 is primarily involved in inhibiting NK cell functions. Two isoforms of human CD244 exist on NK cells, h2B4-A and h2B4-B, which differ in their binding affinity to ligand CD48.
Additional information: Clone REA112 displays negligible binding to Fc receptors.

Alternative names

2B4, NAIL, NKR2B4, Nmrk, SLAMF4

Detailed product information

Technical specifications

CloneREA112
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody (S), human IgG1
Hostcell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Specieshuman
AntigenCD244 (2B4)
Alternative names of antigen2B4, NAIL, NKR2B4, Nmrk, SLAMF4
Molecular mass of antigen [kDa]39
Distribution of antigenNK cells, T cells, eosinophils
Entrez Gene ID51744
RRIDAB_2857608, AB_2857595, AB_2656606, AB_2656607, AB_2656608, AB_2656609, AB_2656610, AB_2656611, AB_2656612, AB_2656613, AB_2656614, AB_2656615, AB_2811678

Resources for CD244 (2B4) Antibody, anti-human, REAfinity™

Certificates

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

References for CD244 (2B4) Antibody, anti-human, REAfinity™

Publications

  1. Munitz, A. et al. (2005) 2B4 (CD244) is expressed and functional on human eosinophils. J. Immunol. 174: 110-118
  2. Sivori, S. et al. (2002) Early expression of triggering receptors and regulatory role of 2B4 in human natural killer cell precursors undergoing in vitro differentiation. Proc. Natl. Acad. Sci. U.S.A. 99: 4526-4531
  3. Mathew, S. O. et al. (2009) Functional role of human NK cell receptor 2B4 (CD244) isoforms. Eur. J. Immunol. 39(6): 1632-1641
  4. Vaidya, S. V. et al. (2006) Of mice and men: different functions of the murine and human 2B4 (CD244) receptor on NK cells. Immunol. Lett. 105(2): 180-184

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