CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

Name: CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™
Availability: InStock

CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™Extended ValidationRecombinant

Clone:

REA197

Type of antibody:

Primary antibodies, Recombinant antibodies

Isotype:

recombinant human IgG1

Applications:

FC, MICS, IF, IHC, 3D-IF

Alternative names:

Siglec-1, SN, Sialic acid-binding Ig-like lectin 1 (Siglec-1)

Flow cytometry applications forCD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

APC
Biotin
PE

Conjugate:

APC

Recommended dilution:

1:50

Remark:

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

Tested:

QC tested

Products used:

130-124-896

Protocols:

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

Description:

Bone marrow cells from BALB/c mice were stained with CD169 (Siglec-1) antibodies or with the corresponding REA control antibodies (left image), as well as with Anti-Gr-1 antibodies and analyzed by flow cytometry using the MACSQuant^® Analyzer. Cell debris and dead cells were excluded from the analysis based on scatter signals and propidium iodide fluorescence.

MICS (MACSima Imaging Cyclic Staining) applications forCD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

APC
PE

Conjugate:

APC

Recommended dilution:

1:50

Fixation:

PFA

Tested:

during development

Species tested:

mouse

Products used:

130-124-896

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

Conjugate:

APC

Recommended dilution:

1:50

Fixation:

Acetone

Tested:

during development

Species tested:

mouse

Products used:

130-124-896

Compatible applications:

IF, IHC

Protocols:

Protocol for MICS experiments

3D-Immunofluorescence applications forCD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

Vio R667

Conjugate:

Vio R667

Recommended dilution:

1:50

Tested:

during development

Products used:

130-131-140

Tissue clearing:

MACS^® Clearing Kit

Compatible applications:

IF, IHC

Description:

3D-immunofluorescence analysis of mouse lymph node fixed with 4% PFA and subjected to whole-mount immunostaining with CD169 Antibody, anti-mouse, Vio R667 (red) and CD31 Antibody, anti-mouse, Vio-570 (yellow). The sample was processed and optically cleared using the MACS^® Clearing Kit and imaged with an UltraMicroscope Blaze light sheet instrument. Sample detail image (A), overview image (B), and 3D-rendered MIP (C). Scale bars: 200 µm (A), 500 µm (B).

Extended validation for CD169 (Siglec-1) Antibody, anti-mouse, 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 REA197
3D6.112	++
Ser-4	-
645608	-
Cells were incubated with an excess of purified unconjugated CD169 (Siglec-1) (REA197) 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 CD169 (Siglec-1). Bone marrow cells from C57BL/6 mice were stained with CD169 (Siglec-1) antibodies and with a suitable counterstaining. As a control, CD169 (Siglec-1) 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 CD169 (Siglec-1). Bone marrow cells from C57BL/6 mice were stained with CD169 (Siglec-1) antibodies and with a suitable counterstaining. As a control, CD169 (Siglec-1) 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 CD169 (Siglec-1). Bone marrow cells from C57BL/6 mice were stained with CD169 (Siglec-1) antibodies and with a suitable counterstaining. As a control, CD169 (Siglec-1) 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 CD169 (Siglec-1). Bone marrow cells from C57BL/6 mice were stained with CD169 (Siglec-1) antibodies and with a suitable counterstaining. As a control, CD169 (Siglec-1) 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 CD169 (Siglec-1) (REA197). 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 CD169 (Siglec-1) (REA197). 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 CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

Overview

Clone REA197 recognizes the mouse CD169 antigen, a Single-pass type I membrane protein also known as Sialoadhesin or Sialic acid-binding Ig-like lectin 1 (Siglec-1). CD169 is a macrophage-restricted cell surface receptor that mediates sialic-acid dependent binding to lymphocytes, including granulocytes, monocytes, natural killer cells, B-cells and CD8

⁺

T-cells. It is a member of the sialic acid-binding IgG-like lectin (Siglec) family of proteins characterized by affinity to specifically sialylated ligands, and under normal conditions is expressed on subsets of macrophages in secondary lymphoid tissues, such as lymph node and spleen. However, CD169

⁺

macrophages can also be found in a variety of pathological conditions, including (autoimmune) inflammatory infiltrates and tumors. CD169 has been shown to contribute to sialylated pathogen uptake, antigen presentation and lymphocyte proliferation, and to influence both immunity and tolerance. Additionally, CD169 positive macrophages, along with mesenchymal stem cells and β-adrenergic neurons, form the hematopoietic stem cell niche in the bone marrow. CD169

⁺

macrophages mediate signaling between the various cells and seem to promote hematopoietic stem cell retention to the niche.

Additional information: Clone REA197 displays negligible binding to Fc receptors.

Alternative names

Siglec-1, SN, Sialic acid-binding Ig-like lectin 1 (Siglec-1)

Detailed product information

Technical specifications

Clone	REA197
Clonality	monoclonal
Isotype	recombinant human IgG1
Isotype control	REA Control Antibody, human IgG1
Host	human cell line
Type of antibody	Primary antibodies, Recombinant antibodies
Species	mouse
Antigen	CD169 (Siglec-1)
Alternative names of antigen	Siglec-1, SN, Sialic acid-binding Ig-like lectin 1 (Siglec-1)
Molecular mass of antigen [kDa]	181
Distribution of antigen	macrophages, spleen
Entrez Gene ID	20612
RRID	AB_2819702, AB_2889469, AB_2928278, AB_2819759

Resources for CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

Certificates

Please follow this

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

References for CD169 (Siglec-1) Antibody, anti-mouse, REAfinity™

Publications

Saunderson, S. C. et al. (2014) CD169 mediates the capture of exosomes in spleen and lymph node. Blood 123(2): 208-216
Chow, A. et al. (2011) Bone marrow CD169+ macrophages promote the retention of hematopoietic stem and progenitor cells in the mesenchymal stem cell niche. J. Exp. Med. 208(2): 261-271
Crocker, P. R. et al. (1994) Sialoadhesin, a macrophage sialic acid binding receptor for haemopoietic cells with 17 immunoglobulin-like domains. EMBO J. 13(19): 4490-4503

Applications

Extended validation