Clone:
REA231
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
FC, MICS, IF, IHC
Alternative names:
CD72b, Lyb-2

Extended validation for CD72 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 REA231
3F3++
J4/117++
Cells were incubated with an excess of purified unconjugated CD72 (REA231) 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.
View details
Overlay histogram showing flow cytometric analysis of CD72 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD72-PE, clone (REA231). 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 CD72 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD72-PE, clone (REA231). 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 CD72. Human peripheral blood mononuclear cells (PBMCs) were stained with CD72 antibodies and with a suitable counterstaining. As a control, CD72 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 CD72. Human peripheral blood mononuclear cells (PBMCs) were stained with CD72 antibodies and with a suitable counterstaining. As a control, CD72 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 CD72. Human peripheral blood mononuclear cells (PBMCs) were stained with CD72 antibodies and with a suitable counterstaining. As a control, CD72 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 CD72. Human peripheral blood mononuclear cells (PBMCs) were stained with CD72 antibodies and with a suitable counterstaining. As a control, CD72 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 CD72. Human peripheral blood mononuclear cells (PBMCs) were stained with CD72 antibodies and with a suitable counterstaining. As a control, CD72 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 CD72 (REA231). 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 CD72 (REA231). 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 CD72 (REA231). 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 CD72 Antibody, anti-human, REAfinity™

Overview

Clone REA231 recognizes CD72, a 45 kDa type II membrane protein with a C-type lectin-like domain. Expression of CD72 is found on most B-lineage cells, follicular dendritic cells, mast cells, and liver Kupffer cells. CD72 is recognized as negative regulator of B cell receptor (BCR) signaling, as well as regulating mast cell growth and differentiation via KIT activation. The inhibitory role of CD72 is exerted via the ITIM sequence in the cytoplasmic tail.
Additional information: Clone REA231 displays negligible binding to Fc receptors.

Alternative names

CD72b, Lyb-2

Detailed product information

Technical specifications

CloneREA231
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody (S), human IgG1
Hosthuman cell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Specieshuman
AntigenCD72
Alternative names of antigenCD72b, Lyb-2
Distribution of antigenB cells, mast cells
Entrez Gene ID971
RRIDAB_2659139, AB_2659140, AB_2659141, AB_2659142, AB_2659143, AB_2659144, AB_2659145, AB_2659146, AB_2659147, AB_2659148, AB_2659149, AB_2659150, AB_2659151, AB_2659138

Resources for CD72 Antibody, anti-human, REAfinity™

Certificates

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

References for CD72 Antibody, anti-human, REAfinity™

Publications

  1. Kataoka, T. R. et al. (2010) CD72 negatively regulates KIT-mediated responses in human mast cells. J Immunol 184: 2468-2475
  2. Li, D. H. et al. (2006) CD72 down-modulates BCR-induced signal transduction and diminishes survival in primary mature B lymphocytes. J Immunol 176: 5321-5328

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