CD16 Antibody, anti-human, REAfinity™

Name: CD16 Antibody, anti-human, REAfinity™
Availability: InStock

CD16 Antibody, anti-human, REAfinity™Extended ValidationRecombinant

Clone:

REA423

Type of antibody:

Primary antibodies, Recombinant antibodies

Isotype:

recombinant human IgG1

Applications:

FC, MICS, IF, IHC, MC

Alternative names:

FCGR3A, FCGR3B, FcR-10, IGFR3, IMD20, FcRIII

Flow cytometry applications forCD16 Antibody, anti-human, REAfinity™

APC
APC-Vio 770
Biotin
FITC
PE
PE-Vio 615
PE-Vio 770
PerCP-Vio 700
Vio Bright B515
Vio Bright R720
Vio Bright V600
VioBlue
VioGreen

Conjugate:

APC

Recommended dilution:

1:50

Remark:

Cells should be stained prior to fixation, if formaldehyde is used as a fixative.

Tested:

QC tested

Reported:

30400618,

Products used:

130-113-951, 130-113-389

Protocols:

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

Description:

Human peripheral blood cells after erythrocyte lysis were stained with CD16 antibodies or with the corresponding REA Control (S) antibodies (left images) as well as with CD15 antibodies. Flow cytometry was performed 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 forCD16 Antibody, anti-human, REAfinity™

PE

Conjugate:

PE

Recommended dilution:

1:50

Fixation:

Acetone

Tested:

during development

Species tested:

human

Products used:

130-113-955, 130-113-393

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

Mass cytometry applications forCD16 Antibody, anti-human, REAfinity™

pure

Conjugate:

pure

Reported:

32676612,

Products used:

130-108-027

Extended validation for CD16 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 REA423
3G8	++
eBioCB16 (CB16)	++
B73.1	++
CLB-Fc-gran1/5D2	++
Cells were incubated with an excess of purified unconjugated CD16 (REA423) 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 CD16. Human peripheral blood mononuclear cells (PBMCs) after erythrocyte lysis were stained with CD16 antibodies and with a suitable counterstaining. As a control, CD16 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 CD16. Human peripheral blood mononuclear cells (PBMCs) after erythrocyte lysis were stained with CD16 antibodies and with a suitable counterstaining. As a control, CD16 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 CD16. Human peripheral blood mononuclear cells (PBMCs) after erythrocyte lysis were stained with CD16 antibodies and with a suitable counterstaining. As a control, CD16 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 CD16. Human peripheral blood mononuclear cells (PBMCs) after erythrocyte lysis were stained with CD16 antibodies and with a suitable counterstaining. As a control, CD16 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 CD16. Human peripheral blood mononuclear cells (PBMCs) after erythrocyte lysis were stained with CD16 antibodies and with a suitable counterstaining. As a control, CD16 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 CD16 (REA423). 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 CD16 (REA423). 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 CD16 Antibody, anti-human, REAfinity™

Overview

Clone REA423 recognizes the human CD16 antigen, a single-pass type I membrane protein which is also known as low affinity immunoglobulin gamma Fc region receptor III (FcRIII). CD16 is a 50–80 kDa glycoprotein that is expressed in two different isoforms. The transmembrane form is found on human NK cells, macrophages, and mast cells, while the glycosylphosphatidylinositol (GPI)-linked form is present on neutrophils. The human CD16 antigen is a low-affinity receptor for aggregated IgG. The transmembrane form plays a role in signal transduction, NK cell activation, and antibody-dependent cellular cytotoxicity. Clone REA423 recognizes both the extracellular domain of the transmembrane form as well as the GPI-linked form of the human CD16 antigen. CD16 is expressed on the majority of rhesus monkey NK cells and on a subset of monocytes but not on granulocytes.

Additional information: Clone REA423 displays negligible binding to Fc receptors.

Alternative names

FCGR3A, FCGR3B, FcR-10, IGFR3, IMD20, FcRIII

Detailed product information

Technical specifications

Clone	REA423
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, non-human primate
Cross-reactivity	rhesus monkey ( Macaca mulatta ) , cynomolgus monkey ( Macaca fascicularis ) , capuchin monkey, chimpanzee ( Pan troglodytes ) , common marmoset ( Callithrix jacchus ) , cotton-top tamarin ( Saguinus oedipus ) , olive baboon ( Papio anubis ) , pigtail monkey ( Macaca nemestrina ) , squirrel monkey ( Saimiri sciureus )
Antigen	CD16
Alternative names of antigen	FCGR3A, FCGR3B, FcR-10, IGFR3, IMD20, FcRIII
Molecular mass of antigen [kDa]	48(sum of molecular weights of subunits)
Distribution of antigen	NK cells, monocytes, granulocytes
Entrez Gene ID	2214, 2215
RRID	AB_2726150, AB_2726428, AB_2726151, AB_2726426, AB_2726149, AB_2726431, AB_2726154, AB_2726432, AB_2726155, AB_2726429, AB_2726152, AB_2733100, AB_2733101, AB_2726430, AB_2726153, AB_2751177, AB_2751110, AB_2751814, AB_2751758, AB_2751968, AB_2751967, AB_2655423, AB_2904778, AB_2904777, AB_2726427

Resources for CD16 Antibody, anti-human, REAfinity™

Documents and Protocols

Brochures/posters

REAfinity™ Antibodies for mass cytometry - Recombinantly engineered to increase reproducibility

Scientific posters

Multicolor immunophenotyping of human tumor cells using MACSQuant® Analyzer 16

App notes/customer reports

Comprehensive flow cytometry Analysis of isolated NK cells from whole blood.

Certificates

Please follow this

link

to search for Certificates of Analysis (CoA) by lot number.

Reviews for CD16 Antibody, anti-human, REAfinity™

CD16 Expression for the Analysis of Human Peripheral Blood Neutrophils

1
2
3
4
5

CD16-FITC, human (130-113-392)

Specifc and bright signal without elaborate optimisation! A great product!

Staining Peripheral Blood with Anti-CD16 PE

1
2
3
4
5

CD16-PE, human (130-113-393)

Works well. Reproducible results.

Staining Peripheral Blood with Anti-CD16 PE

1
2
3
4
5

CD16-PE, human (130-113-955)

Works well. Reproducible results.

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References for CD16 Antibody, anti-human, REAfinity™

Publications

Romee, R. et al. (2013) NK cell CD16 surface expression and function is regulated by a disintegrin and metalloprotease-17 (ADAM17). Blood 121(18): 3599-3608
Ravetch, J. V. et al. (1989) Alternative membrane forms of Fc gamma RIII(CD16) on human natural killer cells and neutrophils. Cell type-specific expression of two genes that differ in single nucleotide substitutions. J. Exp. Med. 170(2): 481-497
Ziegler-Heitbrock, L. (2007)
The CD14
⁺
CD16
⁺
blood monocytes: their role in infection and inflammation.
J. Leukoc. Biol. 584(592): 584-592
Croci, S. et al. (2018) Higher frequencies of lymphocytes expressing the natural killer group 2D receptor in patients with Behçet disease. Front Immunol 9: 2157
Veluchamy, J. et al. (2017)
In vivo
efficacy of umbilical cord blood stem cell-derived NK cells in the treatment of metastatic colorectal cancer.
Front Immunol 8: 87

Applications

Extended validation