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  • 1
    Online Resource
    Online Resource
    Peripheral blood CD115 (CSF-1R) expression is anticoagulant- and time-dependent
    The American Association of Immunologists ; 2019
    In:  The Journal of Immunology Vol. 202, No. 1_Supplement ( 2019-05-01), p. 52.20-52.20
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 202, No. 1_Supplement ( 2019-05-01), p. 52.20-52.20
    Abstract: Human CD115, also known as CSF-1R, is highly expressed by monocytes, macrophages, and dendritic cells. Here we show that detection of human CD115 on peripheral blood monocytes is dependent on both the anticoagulant and the storage time before sample processing. Commercially available CD115 antibody, clone 9-4D2-1E4, shows bright staining on blood samples collected in EDTA and citrate based anticoagulants and a dimmer staining intensity in blood collected in heparin. Clone 9-4D2-1E4 is not compatible with Cyto-Chex™ tubes. Consistent with published reports in mice, human CD115 signal is reduced within hours of collection. CD115 expression is further reduced after overnight storage at ambient temperature and is completely lost two days post collection. Reduced CD115 expression over time is most noticeable in whole blood collected in heparin. These results demonstrate that anti-coagulant and storage time are important considerations for current research projects and clinical trials targeting CD115 (CSF-1R).
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.202.Supp.52.20
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2019
    detail.hit.zdb_id: 1475085-5
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  • 2
    Online Resource
    Online Resource
    A new antibody for the study of human CD157 expression and function
    The American Association of Immunologists ; 2023
    In:  The Journal of Immunology Vol. 210, No. 1_Supplement ( 2023-05-01), p. 79.16-79.16
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 210, No. 1_Supplement ( 2023-05-01), p. 79.16-79.16
    Abstract: CD157 is an ectoenzyme that has both cyclic ADP-ribose hydrolase and ADP-ribosyl cyclase activities. In the human hematopoietic system, CD157 is prevalently expressed by cells of the myelomonocytic lineage, and interacts with extracellular matrix components and regulates leukocyte diapedesis via integrin-mediated signaling. CD157 is implicated in the pathophysiology of several neurological disorders. Although it lacks a cytoplasmatic domain, CD157 is able to transduce intracellular signals by establishing functional and structural crosstalk with β1 (CD29) and β2 (CD18) integrins. CD157 ligation causes integrin– dependent cytoskeletal remodeling and increase in F-actin content. Activation of downstream signaling pathways leads to cell polarization, with CD157 migrating prevalently to the rear of the cells whereas F-actin is localized at the opposite pole. However, the molecular and cellular functions of CD157 still remain poorly understood. Here, we describe the generation of a monoclonal antibody specific for human CD157 (clone W21007F) that can be used in flow cytometry, immunofluorescence and blocking of CD157 mediated adhesion of monocytes to fibronectin. Treatment of the pro-monocytic, myeloid leukemia cell line U937 with clone W21007F induces cell polarization and a marked increase in filamentous actin, with CD157 clustering at the uropod. The majority of F-Actin fibers are located at the opposite end of CD157, and CD157 co-localizes with integrin beta-1 in distinct membrane domains. The reference clone SY/11B5 and isotype control antibody do not show this effect. In conclusion, anti-human CD157 (clone W21007F) is a valuable tool for the study of CD157 function and expression in human cells.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.210.Supp.79.16
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2023
    detail.hit.zdb_id: 1475085-5
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  • 3
    Online Resource
    Online Resource
    Novel monoclonal antibodies for the detection of human TLR9
    The American Association of Immunologists ; 2019
    In:  The Journal of Immunology Vol. 202, No. 1_Supplement ( 2019-05-01), p. 64.15-64.15
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 202, No. 1_Supplement ( 2019-05-01), p. 64.15-64.15
    Abstract: Human TLR9 (CD289) is a well characterized mediator of antiviral immunity. TLR9 is primarily activated by unmethylated CpG sequences leading to the production of type I interferons and other inflammatory cytokines. Recently, changes in TLR9 expression in cancer have also been reported. Here we describe novel monocloncal antibodies for the detection of human TLR9 by flow cytometry. These clones recognize intracellular TLR9 in B cells and plasmacytoid dendritic cells (pDCs), as well as on HEK293 cells transfected with human TLR9, and are compatible with multiple fixation and permeabilization buffers. We also show surface expression of TLR9 on pDCs, which is reported to be upregulated following stimulation. In response to CpG-B stimulation, two clones enhance and inhibit TLR9 signaling in a concentration dependent manner leading to alterations in cytokine secretion. Reported here are multiple human TLR9 clones with greater specificity than other commercially available clones.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.202.Supp.64.15
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2019
    detail.hit.zdb_id: 1475085-5
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  • 4
    Online Resource
    Online Resource
    Role of C-type lectin receptor CLEC12A in human basophil activation (48.6)
    The American Association of Immunologists ; 2012
    In:  The Journal of Immunology Vol. 188, No. 1_Supplement ( 2012-05-01), p. 48.6-48.6
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 188, No. 1_Supplement ( 2012-05-01), p. 48.6-48.6
    Abstract: CLEC12A is a member of the C-type lectin receptor family with a cytoplasmic immunoreceptor tyrosine-based inhibitory motif. Murine CLEC12A is expressed on variety of cell types including dendritic cells, monocytes/macrophages, B cells, neutrophils and basophils but not NK cells. In humans, CLEC12A is reported to be expressed on dendritic cells (DC, both myeloid and plasmacytoid), monocytes, B cells (normal and leukemic) and NK cells, but not on T cells or basophils. Performing flow cytometric analysis of human peripheral blood leukocytes using an anti-human CLEC12A antibody (clone 50C1), we report CLEC12A expression on a subset of CD3, CD19 and CD56 negative cells. Further analysis revealed these cells to be CCR3+ basophils. Antibody mediated ligation of CLEC12A on DCs results in cytokine production and cellular maturation and can inhibit NK cell mediated cytotoxicity. In response to IgE and IL-3, basophils can induce Th2 differentiation and amphiregulin production, respectively. We hypothesize that ligation with CLEC12A will alter IL-3 and/or IgE mediated basophil activation, which will be studied in our ongoing work.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.188.Supp.48.6
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2012
    detail.hit.zdb_id: 1475085-5
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  • 5
    Online Resource
    Online Resource
    Discrimination of spectrally overlapping fluorochromes using Sony Corporations’s spectral analyzer (SP6800) (TECH1P.865)
    The American Association of Immunologists ; 2014
    In:  The Journal of Immunology Vol. 192, No. 1_Supplement ( 2014-05-01), p. 69.33-69.33
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 192, No. 1_Supplement ( 2014-05-01), p. 69.33-69.33
    Abstract: A maximum of seven colors can be separated using traditional 2-laser flow cytometers and it is required to have the necessary band pass filters to separate these fluorochromes. In addition, these systems cannot differentiate between spectrally overlapping fluorophores such as PerCP and PerCP/Cy5.5; or FITC, PE and Alexa Fluor 514® (A514). SP6800, a novel spectral analyzer from Sony Corporation uses an innovative optics system that collects all emitted light and eliminates the need for band pass filters. We performed whole blood staining using a panel of 9 markers, which included significantly overlapping fluorochromes PerCP and PerCP/Cy5.5 and A514, PE and FITC and analyzed it on the SP6800. We were able clearly identify CD56+ (PerCP/Cy5.5), CD19+ (PerCP) and CD3+ (A514) cells. Further comparison of a 7-color panel of consisting of CD3 FITC, CD56 PE, CD19 PerCP, CD8 PE/Cy7, CD16 APC, CD4 Alexa Fluor 700® and CD62L APC/Cy7 on the BD LSRFortessa™ system and SP6800 gave similar results. Our data show that the SP6800 can perform all the analysis similar to a traditional cytometer with the added feature of separating spectrally overlapping fluorophores.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.192.Supp.69.33
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2014
    detail.hit.zdb_id: 1475085-5
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  • 6
    Online Resource
    Online Resource
    Standardized oligo barcode antibody conjugates, and Veri-Cells™ PBMC for multiplex immunophenotyping and CITE-Seq
    The American Association of Immunologists ; 2018
    In:  The Journal of Immunology Vol. 200, No. 1_Supplement ( 2018-05-01), p. 174.19-174.19
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 200, No. 1_Supplement ( 2018-05-01), p. 174.19-174.19
    Abstract: The CITE-Seq (Cellular Indexing of Transcriptomes and Epitopes by Sequencing) platform is a recent advance in single cell analysis, which is based on high-throughput single cell sequencing (scSeq) and combines measurements of cellular proteins and transcriptomes. This platform will potentially transform how complex cell populations (lineage differentiation or tumor infiltrating lymphocytes) are studied. Published data indicated that CITE-Seq analysis on cell surface marker expression are comparable to multi-color flow cytometry, but provides superior capacity in multiplexing to the latter. Currently, individual investigators use their choice of oligo bar-codes for different protein markers and there is no available control. This will make comparison of data from different studies difficult in the future. The availability of standardized oligonucleotide bar-code labeled antibodies as well as control cells will enable reliable comparison data from longitudinal studies or among different studies. FlexSeq™ is our new product line, in which monoclonal antibodies are conjugated with unique oligonucleotide bar-codes. We provide standardized bar-coding system and ready to use oligo-antibody conjugates to support CITE-Seq based multiplex immunophenotyping. Veri-Cells™ is our proprietary lyophilized cell product that can be used as a control for detection of cell surface and intracellular flow cytometry. In this study, we validate the application of FlexSeq™ products in CITE-Seq platform, and compare Veri-Cells™ PBMC with freshly isolated PBMC in the assays.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.200.Supp.174.19
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2018
    detail.hit.zdb_id: 1475085-5
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  • 7
    Online Resource
    Online Resource
    Expression of Ikaros family member molecules in lupus-prone mice: a preliminary analysis (P4102)
    The American Association of Immunologists ; 2013
    In:  The Journal of Immunology Vol. 190, No. 1_Supplement ( 2013-05-01), p. 51.17-51.17
    Details
    In: The Journal of Immunology, The American Association of Immunologists, Vol. 190, No. 1_Supplement ( 2013-05-01), p. 51.17-51.17
    Abstract: Ikaros family of zinc-finger proteins, namely Aiolos, Helios and Ikaros, are hematopoietic-specific transcription factors involved in lymphocyte development. Aiolos-deficient mice develop SLE-like disease, and Ikaros family members have been linked to the balance between T cell activation and tolerance. To begin to investigate the role of Ikaros proteins in SLE, we first determined the expression of these proteins in spleen cells of NZM.2328 mice that develop disease that mimics human SLE in many ways, including a profound female bias. Results show that Aiolos expression in T cells is significantly lower than that in B cells from NZM.2328 mice. Similar reduction in Aiolos expressing T cells was observed in old lupus-prone MRL-MpJ-Fas+/+ compared to age-matched control B6 mice. Almost all B cells expressed Aiolos in all mice tested, but the expression level was much lower in MRL-MpJ+/+ B cells than in control B6 B cells. In contrast to Aiolos expression, T cells expressing Helios were 3-4-fold higher in old NZM.2328 females than in age-matched male and younger female mice. Ikaros expression was also higher on both T and B cells in older NZM.2328 females than in age-matched male and younger female mice. Ongoing studies will investigate the relationship of dysregulated Ikaros family molecules with lupus disease and examine the role of this dysregulation in female bias and lupus pathogenesis.
    Type of Medium: Online Resource
    ISSN: 0022-1767 , 1550-6606
    URL: Article
    DOI: 10.4049/jimmunol.190.Supp.51.17
    RVK:
    XA 54100
    RVK:
    XA 10000
    Language: English
    Publisher: The American Association of Immunologists
    Publication Date: 2013
    detail.hit.zdb_id: 1475085-5
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