Clone:
REAL258
Type of antibody:
Releasable antibodies, Primary antibodies, Recombinant antibodies
Applications:
FC, MICS, IHC, IF
Alternative names:
Ptprc, GP180, L-CA, LY5, T200

Extended validation for CD45 Antibody, anti-human,
REAlease
®

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 REAL258
5B1++
2D1++
HI30++
MB4-6D6-
REA1023-
REA747++
Cells were incubated with an excess of purified unconjugated CD45 (REAL258) 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 CD45 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45-PE (REAL258, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Fluorescence microscopy image of CD45 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45-PE (REAL258, red) and counterstained with DRAQ5 (blue) as DNA stain.
Fluorescence microscopy image of CD45 knockout cells. Wild type (WT, left) and knockout cells (KO, right) were stained with CD45-PE (REAL258, red) and counterstained with DRAQ5 (blue) as DNA stain.
View details
Overlay histogram showing flow cytometric analysis of human CD45 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD45-PE, clone (REAL258). 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 CD45 knockout cells. Wild type (red) and knockout cells (blue) were stained with CD45-PE, clone (REAL258). 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.

Specifications for CD45 Antibody, anti-human,
REAlease
®

Overview

Clone REAL258 is an antibody fragment derived from the full CD45 antibody molecule. It displays no binding to Fc receptors. The recombinantly engineered antibody fragments are multimerized to form the REAlease Complex to bind markers with high avidity.
Clone REAL258 recognizes the human CD45 antigen, a member of the protein tyrosine phosphatase (PTP) family, also known as the leukocyte common antigen (LCA). The CD45 molecule is a type I transmembrane protein which is required for T and B cell activation and involved in cell growth, differentiation, mitotic cycle, and oncogenic transformation. CD45 is expressed in different isoforms (CD45RA, CD45RB, CD45RC, CD45RAB, CD45RAC, CD45RBC, CD45RO, CD45R (ABC)) depending on the differentiation status of the cell.
The REAlease Kits consist of the respective fluorochrome-conjugated REAlease Complexes and the REAlease Support Kit for removal of the REAlease Complexes and optional relabeling with different fluorochrome-conjugated REAlease Complexes.

Alternative names

Ptprc, GP180, L-CA, LY5, T200

Detailed product information

Technical specifications

CloneREAL258
Clonalitymonoclonal
Isotype controlControl Antibody
Hostcell line
Type of antibodyReleasable antibodies, Primary antibodies, Recombinant antibodies
Specieshuman
AntigenCD45
Alternative names of antigenPtprc, GP180, L-CA, LY5, T200
Distribution of antigenT cells, B cells, leukocytes
RRIDAB_2751276, AB_2751434, AB_2751416, AB_2751805, AB_2751745, AB_2751288, AB_2751280, AB_2801811, AB_2801804, AB_2784199, AB_2784200, AB_2784198, AB_2784201, AB_2811324, AB_2811707, AB_2751284

References for CD45 Antibody, anti-human,
REAlease
®

Publications

  1. Ralph, S. J. et al. (1987) Structural variants of human T200 glycoprotein (leukocyte-common antigen). EMBO J. 6(5): 1251-1257
  2. Vykoukal, J. et al. (2008) Enrichment of putative stem cells from adipose tissue using dielectrophoretic field-flow fractionation. Lab Chip 8(8): 1386-1393
  3. Kurian, L. et al. (2013) Conversion of human fibroblasts to angioblast-like progenitor cells. Nat. Methods 10(1): 77-83

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