CD52 Antibody, anti-human, REAfinity™

Name: CD52 Antibody, anti-human, REAfinity™
Availability: InStock

CD52 Antibody, anti-human, REAfinity™Extended ValidationRecombinant

Clone:

REA164

Type of antibody:

Primary antibodies, Recombinant antibodies

Isotype:

recombinant human IgG1

Applications:

FC, MICS, IF, IHC

Alternative names:

CDW52, CAMPATH-1

Flow cytometry applications forCD52 Antibody, anti-human, REAfinity™

APC-Vio 770
Biotin
FITC
PE
PE-Vio 615
PE-Vio 770
PerCP-Vio 700
Vio Bright B515
Vio Bright R720
VioBlue

Conjugate:

APC-Vio 770

Recommended dilution:

1:50

Remark:

The antibody is suited for staining of formaldehyde-fixed cells.

Tested:

QC tested

Products used:

130-130-067, 130-129-979

Protocols:

Cell surface flow cytometry staining protocol (PBS/EDTA/BSA 1:50)

Description:

Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies as well as with CD3 antibodies and analyzed by flow cytometry using the MACSQuant^® Analyzer. The Tandem Signal Enhancer has been used to increase binding specificity of tandem-dye–conjugated antibodies. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide fluorescence or 4',6-diamidino-2-phenylindole (DAPI) fluorescence, as in the case of tandem conjugates.

Conjugate:

APC-Vio 770

Recommended dilution:

1:11

Remark:

The antibody is suited for staining of formaldehyde-fixed cells.

Tested:

QC tested

Products used:

130-099-617, 130-099-619

Protocols:

Cell surface flow cytometry staining protocol (PBS/EDTA/BSA 1:11)

Description:

Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies as well as with CD3 antibodies and analyzed by flow cytometry using the MACSQuant^® Analyzer. The Tandem Signal Enhancer has been used to increase binding specificity of tandem-dye–conjugated antibodies. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide fluorescence or 4',6-diamidino-2-phenylindole (DAPI) fluorescence, as in the case of tandem conjugates.

MICS (MACSima Imaging Cyclic Staining) applications forCD52 Antibody, anti-human, REAfinity™

FITC
PE

Conjugate:

FITC

Recommended dilution:

1:50

Fixation:

FFPE

Tested:

during development

Species tested:

human

Products used:

130-123-926, 130-123-680

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

Conjugate:

FITC

Recommended dilution:

1:50

Fixation:

PFA

Tested:

during development

Species tested:

human

Products used:

130-123-926, 130-123-680

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

Conjugate:

FITC

Recommended dilution:

1:50

Fixation:

Acetone

Tested:

during development

Species tested:

human

Products used:

130-123-926, 130-123-680

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

Extended validation for CD52 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 clones	Overlap in epitope recognition with REA164
4C8	+
Hu116 (Campath-1H)	++
QA19A22	++
YTH34.5	++
Cells were incubated with an excess of purified unconjugated CD52 (REA164) 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.

Sensitivity

Performance comparison

Selected fluorochrome conjugated antibodies from Miltenyi Biotec were compared to commercially available hybridoma clones in flow cytometry analysis.

Flow cytometric comparison of different clones for CD52. Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies and with a suitable counterstaining. As a control, CD52 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 CD52. Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies and with a suitable counterstaining. As a control, CD52 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 CD52. Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies and with a suitable counterstaining. As a control, CD52 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 CD52. Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies and with a suitable counterstaining. As a control, CD52 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 CD52. Human peripheral blood mononuclear cells (PBMCs) were stained with CD52 antibodies and with a suitable counterstaining. As a control, CD52 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

Comparison of staining pattern on non-fixed and fixed cells using CD52 (REA164). 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 CD52 (REA164). 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 CD52 Antibody, anti-human, REAfinity™

Overview

Clone REA164 recognizes CD52, a glycosylphosphatidylinositol (GPI)-anchored, 25–29 kDa heavily glycosylated antigen. The CD52 core peptide is only 12 amino acids long and is linked to the surface via C-terminal GPI anchor. Expression of CD52 is found on lymphocytes, monocytes, macrophages, eosinophils, and in the male reproductive tract on the epithelial cells of the epididymis and seminal vesicle. CD52 is absent in immature spermatozoa in the efferent ducts but is acquired by the sperm membrane, progressively, as they pass through the epididymis proper, where CD52 is shed into the seminal plasma by the epididymis. Though the exact role of CD52 is unclear, antibodies against CD52 elicit antibody dependent cellular and complement mediated cytolysis of the CD52 expressing lymphocytes. Owing to this, such antibodies are often used to target malignant cells. CD52 expressed on sperms and on lymphocytes are basically different glycoforms of the same core peptide.

Additional information: Clone REA164 displays negligible binding to Fc receptors.

Alternative names

CDW52, CAMPATH-1

Detailed product information

Technical specifications

Clone	REA164
Clonality	monoclonal
Isotype	recombinant human IgG1
Isotype control	REA Control Antibody (S), human IgG1
Host	human cell line
Type of antibody	Primary antibodies, Recombinant antibodies
Species	human
Antigen	CD52
Alternative names of antigen	CDW52, CAMPATH-1
Molecular mass of antigen [kDa]	1
Distribution of antigen	lymphocytes
Entrez Gene ID	1043
RRID	AB_2819508, AB_2889602, AB_2889601, AB_2658646, AB_2658647, AB_2658648, AB_2658649, AB_2658650, AB_2658651, AB_2658652, AB_2658653, AB_2658654, AB_2658655, AB_2921961, AB_2921941, AB_2905071, AB_2905070, AB_2928509, AB_2928508, AB_2928511, AB_2928510, AB_2928513, AB_2928512, AB_2819543

Resources for CD52 Antibody, anti-human, REAfinity™

Certificates

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References for CD52 Antibody, anti-human, REAfinity™

Publications

Ambrose, L. R. et al. (2009) Neutrophils express CD52 and exhibit complement-mediated lysis in the presence of alemtuzumab. Blood 114: 3052-3055
Rawstron, A. C. et al. (2006) CD52 expression patterns in myeloma and the applicability of alemtuzumab therapy. Haematologica 91: 1577 -1578
Yeung, C. H. et al. (1998) Epididymal secretion of CD52 as measured in human seminal plasma by a fluorescence immunoassay. Mol. Hum. Reprod. 4: 447-451

Applications

Extended validation