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>   home   >   Products   >   Primary Antibodies   >   Microbiology   >   Anti-Human CD169 Antibody, clone 7-239 (RPE)   

Anti-Human CD169 Antibody, clone 7-239

Mouse Anti-Human Monoclonal Antibody

     
  • FC - Anti-Human CD169 Antibody, clone 7-239  ABD12296
    Published customer image:AlexaFluor488 conjugated Mouse anti Human CD169 antibody, clone 7-239 used for the detection of CD169 by flow cytometry.Image caption: Introduction of a di-aromatic motif in CT of CD169 results in endocytosis of HIV-1 particles.(A) THP-1 cells expressing either wild type (WT) CD169 or CT mutant CD169YF were incubated with saturating amounts (10 µg/ml) of anti-human CD169 antibody or corresponding isotype controls for 30 minutes at 4°C. Cells were washed twice at 4°C and then an aliquot (1x105 cells) was removed and placed on ice (time point 0 min). The remaining cells were then shifted to 37°C for 30 minutes to promote endocytosis. To arrest internalization, cells were transferred to 4°C, and the number of antibody-bound CD169 molecules left at the surface revealed by staining with PE-conjugated goat anti-mouse IgG antibody and analyzed by flow cytometry. The mean fluorescence intensity (MFI) of the isotype controls was subtracted at each time points and MFIs at 30 min were normalized to that observed at 0 min. The data shown is the percent of anti-CD169 antibody remaining at the cell surface 30 minutes post incubation at 37°C and is the mean ± SEM of four independent experiments. (B) Cells were incubated with Gag-mCherry VLPs and stained for plasma membrane bound CD169 (Surface, top panel) or total CD169 (+ Tx100, bottom panel). CD169 (green), Gag-mCherry VLP (red) and nucleus (blue). Representative deconvolved images of single slices of cells are shown. Scale bars represent 5 µm. (C) Co-localization between green and red (VLPs) signals is reported as mean Pearson’s coefficient ± SEM. Each dot represents a single cell. Two-tailed P values were calculated using unpaired t-test in GraphPad Prism 5. *: P < 0.05, **: P < 0.01.From:Akiyama H, Ramirez N-GP, Gudheti MV, Gummuluru S CD169-Mediated Trafficking of HIV to Plasma Membrane Invaginations in Dendritic Cells Attenuates Efficacy of Anti-gp120 Broadly Neutralizing Antibodies. PLoS Pathog 11: e1004751.
  • WB - Anti-Human CD169 Antibody, clone 7-239  ABD12296
    Published customer image:AlexaFluor488 conjugated Mouse anti Human CD169 antibody, clone 7-239 used for the detection of CD169 by flow cytometry.Image caption:The cytoplasmic tail of CD169 is dispensable for mediating HIV-1 trans-infection. (A) Sequences of wild type and mutant CD169 CTs. The asterisks represent stop codons introduced into the ORFs of the two CT mutants. (B) Western blot analysis of THP-1 cell lysates expressing either wild type or mutant CD169. (C) Cell surface expression of CD169 on THP-1 cells was measured by flow cytometry. (D) Relative cell surface expression of CD169 CT mutants was quantified and normalized to that observed with THP-1/CD169 cells. (E) Cells were challenged with HIV-1, washed and cell-associated p24gag was measured. The data shown is the virus capture by THP-1/CD169 CT mutants (?CT or ?CT4R) normalized to that observed with THP-1/CD169 cells. (F) THP-1/CD169- or THP-1/CD169 CT mutant-mediated trans-infection was determined by measuring luciferase activity in THP—CD4+ T cell co-cultures 2 days post initiation of co-culture. The data shown is the relative virus transmission by THP-1/CD169 CT mutants (?CT or ?CT4R) to that observed with THP-1/CD169 cells. (G) Efficacy of trans-infection was calculated as trans-infection (luciferase activity) per amount of virus captured (cell-associated p24gag) and normalized to that observed with THP-1/CD169 cells (set as 100). The data shown are the means ± SEM of three (D to F) or four (G) independent experiments. (H) THP-1/CD169 or THP-1/CD169?CT4R cells were incubated with Gag-mCherry VLPs , washed, fixed and stained for CD169 and nucleus . Representative deconvolved images of single slices of cells are shown. Scale bar represents 5 µm. WT: THP-1/CD169, ?CT: THP-1/CD169?CT, ?CT4R: THP-1/CD169?CT4R and Vec: empty vector transduced THP-1.From:Akiyama H, Ramirez N-GP, Gudheti MV, Gummuluru S CD169-Mediated Trafficking of HIV to Plasma Membrane Invaginations in Dendritic Cells Attenuates Efficacy of Anti-gp120 Broadly Neutralizing Antibodies. PLoS Pathog 11(3): e1004751.
  • E - Anti-Human CD169 Antibody, clone 7-239  ABD12296
    Published customer image:Mouse anti Human CD169 antibody used for the detection of sialoadhesin on dendritic cells by immunofluorescence.Image caption:Siglec-1 captures HIV-1 and traffics with the virus to the same sac-like compartment.(A) Comparative capture of HIV-1 by distinct DCs that had been pre-incubated with 10 µg/ml of the indicated mAbs or 500 µg/ml of mannan for 30 min before viral exposure. Cells were cultured with HIV-1 in the presence of the indicated reagents, washed, and lysed to measure p24Gag by ELISA. Viral binding at 4°C in LPS mDCs is shown in Figure S3. Data show mean values and SEMs from two experiments including cells from six donors. (B) Comparative capture of HIV-1 by distinct DCs first exposed to the virus and then treated with the indicated reagents for 30 min before washing. Cells were lysed and assessed by p24Gag ELISA. Data show mean values and SEMs from two experiments including cells from six donors. (C) Comparative capture of HIV-1 by distinct blood myeloid DCs that had been pre-incubated with 10 µg/ml of the indicated mAbs for 30 min before viral exposure as in panel A. Figure S4 depicts Siglec-1 surface expression levels of blood myeloid cells. Data show mean values and SEMs from two experiments including cells from six donors. (D) Confocal microscopy analysis of LPS mDCs pulsed for 4 h with GM1-containing LUVHIV-tRed, VLPHIV-Gag-Cherry or HIV-1Cherry, fixed, permeabilized, and then stained for Siglec-1 with mAb 7–239-Alexa 488. Merge of the bright field and maximun fluorescence intensity (scale bar: 5 µm). (3D images) Isosurface representation of DAPI stained nucleus and maximum fluorescence intensity of the sac-like compartment where particles and Siglec-1 accumulate are shown in a 3D volumetric x-y-z data field. (Bar graphs) Quantification of the percentage of GM1-containing LUVHIV-tRed, VLPHIV-Gag-Cherry or HIV-1Cherry co-localizing with Siglec-1-Alexa 488 7–239 and vice versa, obtained analyzing at least 50 compartments from LPS mDCs of two donors. The mean and standard deviation of the thresholded correlation coefficient of Pearson (obtained considering all the images) were 0.77±0.07, indicating co-localization. See also Movies S1, S2, S3 or Figure S5 to observe the compartment in relation to the plasma membrane or the cytoplasm of the cells. (E) Confocal microscopy analysis showing the sac-like compartment pattern of Siglec-1 in LPS mDCs after internalization of the a-Siglec-1 mAb 7D2. Cells were labeled with the mAb for 30 min at 16°C, revealed with an Alexa 488 secondary Ab, shifted to 37°C for 4 h, and analyzed. (3D image) 3D reconstruction (representative of 69% of the analyzed DCs) was done as in (D). Merge of the bright field and maximun fluorescence intensity (scale bar: 5 µm).From: Izquierdo-Useros N, Lorizate M, Puertas MC, Rodriguez-Plata MT, Zangger N, et al. Siglec-1 Is a Novel Dendritic Cell Receptor That Mediates HIV-1 Trans-Infection Through Recognition of Viral Membrane Gangliosides. PLoS Biol 10: e1001448.
  • E - Anti-Human CD169 Antibody, clone 7-239  ABD12296
    Published customer image:Mouse anti Human CD169 antibody used for the detection of sialoadhesin on dendritic cells by flow cytometry.Image caption:Siglec-1 is up-regulated in highly trans-infecting LPS mDCs. (A) Comparative HIV-1 capture of LPS and ITIP mDCs: cells were cultured with HIV-1, washed, and lysed to measure viral p24Gag antigen by ELISA. Comparative transmission of captured HIV-1 from LPS and ITIP mDCs to a reporter CD4+ cell line. Graphs show mean values and standard error of the means from two independent experiments including cells from six donors. (B) Plot of SIGLEC genes (in open circles), CD86 and DC-SIGN (in grey circles) computing the fold change in LPS mDCs compared to ITIP mDCs, and the average gene expression across all samples. Circle size is inversely proportional to adjusted p values. Highlighted in red are statistically differentially expressed genes. Analysis was performed with DCs from four donors matured in parallel with the different stimuli. (C) Relative quantification of SIGLEC1 mRNA expression levels in distinct DCs analyzed by qRT-PCR. Measurements were normalized using the endogenous control housekeeping gene Beta Glucuronidase. Data show means and SEMs of samples from six donors. (D) Cell surface expression of Siglec-1 in distinct DCs analyzed by FACS with mAb 7–239-PE. (Left graph) Geometric mean fluorescence intensity of Siglec-1. (Right graph) Percentage of Siglec-1 positive cells. Data show mean values and SEM from two experiments, including cells from six donors. Representative profiles of Siglec-1 staining in distinct DCs derived from one donor.From: Izquierdo-Useros N, Lorizate M, Puertas MC, Rodriguez-Plata MT, Zangger N, et al. Siglec-1 Is a Novel Dendritic Cell Receptor That Mediates HIV-1 Trans-Infection Through Recognition of Viral Membrane Gangliosides. PLoS Biol 10: e1001448.
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Product Information
Application
  • Applications Legend:
  • WB=Western Blot
  • IHC=Immunohistochemistry
  • IHC-P=Immunohistochemistry (Paraffin-embedded Sections)
  • IHC-F=Immunohistochemistry (Frozen Sections)
  • IF=Immunofluorescence
  • FC=Flow Cytopmetry
  • IC=Immunochemistry
  • ICC=Immunocytochemistry
  • E=ELISA
  • IP=Immunoprecipitation
  • DB=Dot Blot
  • CHIP=Chromatin Immunoprecipitation
  • FA=Fluorescence Assay
  • IEM=Immunoelectronmicroscopy
  • EIA=Enzyme Immunoassay
IHC-F, FC, IP
Primary Accession Q9BZZ2
Reactivity Human
Host Mouse
Clonality Monoclonal
Isotype IgG1
Clone Names 7-239
Calculated MW 182624 Da
Additional Information
Purification Purified IgG prepared by affinity chromatography on Protein G from tissue culture supernatant
Immunogen Human Rhinovirus (HRV14) treated monocyte-derived dendritic cells.
Shelf Life 18 months from date of despatch.
Gene ID 6614
Other Names Sialoadhesin, Sialic acid-binding Ig-like lectin 1, Siglec-1, CD169, SIGLEC1, SN
Target/Specificity Mouse anti-Human CD169 antibody, clone 7-239 specifically recognizes human CD169, otherwise known as Sialoadhesin (Sn), a type I transmembrane glycoprotein and prototypic member of the Siglec (sialic acid binding Ig-like lectin) family, designated Siglec-1. CD169 is a macrophage-restricted interaction molecule which binds to sialylated ligands on haematopoietic cells, including neutrophils, NK cells, monocytes, a subset of CD8+ T cells and B cells, through recognition of sialic acid in the alpha-2,3- glycosidic linkage. CD169 is highly expressed by dendritic cells and on stromal macrophages in the spleen, bone marrow and lymph nodes and to a lesser extent in liver, lungs and gut, and studies have shown high expression of CD169 on macrophages in the inflammatory conditions rheumatoid arthritis and atherosclerosis. Clone 7-239 completely inhibits erythrocyte rosetting around Sn+ transductants.
Preservative & Stabilisers 0.09% Sodium Azide
Storage Store at +4℃ or at -20 ℃.
PrecautionsAnti-Human CD169 Antibody, clone 7-239 is for research use only and not for use in diagnostic or therapeutic procedures.
Research Areas
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References

1. Kirchberger, S. et al. (2005) Human rhinoviruses inhibit the accessory function of dendritic cells by inducing sialoadhesin and B7-H1 expression.
J Immunol. 175 (2): 1145-52. 2. Izquierdo-Useros, N. et al. (2012) Siglec-1 Is a Novel Dendritic Cell Receptor That Mediates HIV-1 Trans-Infection Through Recognition of Viral Membrane Gangliosides.
PLoS Biol. 10: e1001448. 3. Rose, T. et al. (2013) IFNa and its response proteins, IP-10 and SIGLEC-1, are biomarkers of disease activity in systemic lupus erythematosus.
Ann Rheum Dis. 72 (10): 1639-45. 4. Akiyama H et al. (2015) CD169-Mediated Trafficking of HIV to Plasma Membrane Invaginations in Dendritic Cells Attenuates Efficacy of Anti-gp120 Broadly Neutralizing Antibodies.
PLoS Pathog. 11 (3): e1004751.1. Hartnell, A. et al. (2001) Characterization of human sialoadhesin, a sialic acid binding receptor expressed by resident and inflammatory macrophage populations.
Blood. 97 (1): 288-96.

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