Clone:
REA119
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
FC, MICS, IF, IHC
Alternative names:
Ptprc, B220, CD45, CD45R, GP180, L-CA, LCA, LY5, T200

Extended validation for CD45RB 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 REA119
MEM-55++
MT-4-
Cells were incubated with an excess of purified unconjugated CD45RB (REA119) 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 CD45RB knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45RB-PE (REA119, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Fluorescence microscopy image of CD45RB knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45RB-PE (REA119, red) and counterstained with DRAQ5 (blue) as DNA stain.
Fluorescence microscopy image of CD45RB knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45RB-PE (REA119, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Overlay histogram showing flow cytometric analysis of human CD45RB knockout cells. Wild type (red) and knockout cells (blue) were stained with CD45RB-PE, clone (REA119). 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 CD45RB knockout cells. Wild type (red) and knockout cells (blue) were stained with CD45RB-PE, clone (REA119). 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 CD45RB. Human peripheral blood mononuclear cells (PBMCs) were stained with CD45RB antibodies and with a suitable counterstaining. As a control, CD45RB 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 CD45RB. Human peripheral blood mononuclear cells (PBMCs) were stained with CD45RB antibodies and with a suitable counterstaining. As a control, CD45RB 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 CD45RB. Human peripheral blood mononuclear cells (PBMCs) were stained with CD45RB antibodies and with a suitable counterstaining. As a control, CD45RB 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 CD45RB. Human peripheral blood mononuclear cells (PBMCs) were stained with CD45RB antibodies and with a suitable counterstaining. As a control, CD45RB 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 CD45RB (REA119). 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 CD45RB (REA119). 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 CD45RB (REA119). 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 CD45RB Antibody, anti-human, REAfinity™

Overview

Clone REA119 recognizes CD45RB, which is a splice variant isoform of CD45, the leukocyte common antigen (LCA). CD45 is a haemopoietic cell specific protein tyrosine phosphatase, and regulates various signaling events through cytokine receptors and in response to cellular adhesion. Expression of CD45RB is found on T cell subset, B cells, monocytes/macrophages, granulocytes, and dendritic cells.
Additional information: Clone REA119 displays negligible binding to Fc receptors.

Alternative names

Ptprc, B220, CD45, CD45R, GP180, L-CA, LCA, LY5, T200

Detailed product information

Technical specifications

CloneREA119
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody (S), human IgG1
Hostcell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Specieshuman, non-human primate
Cross-reactivity
rhesus monkey (
Macaca mulatta
)
AntigenCD45RB
Alternative names of antigenPtprc, B220, CD45, CD45R, GP180, L-CA, LCA, LY5, T200
Molecular mass of antigen [kDa]145
Distribution of antigenB cells, NK cells, T cells
Entrez Gene ID5788
RRIDAB_2876979, AB_2658345, AB_2658346, AB_2658347, AB_2658348, AB_2658349, AB_2658350, AB_2658351, AB_2658352, AB_2658353, AB_2658354, AB_2658355, AB_2658356, AB_2876978

Resources for CD45RB Antibody, anti-human, REAfinity™

References for CD45RB Antibody, anti-human, REAfinity™

Publications

  1. Parikh, K. et al. (2009) Extracellular ligation-dependent CD45RB enzymatic activity negatively regulates lipid raft signal transduction. Blood 113: 594-603
  2. Dianzani, U. et al. (1992) Isoform-specific associations of CD45 with accessory molecules in human T lymphocytes. Eur. J. Immunol. 22(2): 365-371

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