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BB515 Mouse Anti-Human Siglec-9 (CD329)
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Multiparameter flow cytometric analysis of Siglec-9 (CD329) expression on Human peripheral blood leukocyte populations. Human whole blood was first treated with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899) to lyse erythrocytes, washed, and then preincubated with BD Pharmingen™ Human BD Fc Block™ (Cat. No. 564219/564220). The leukocytes were then stained with either BD Horizon™ BB515 Mouse IgG1, κ Isotype Control (Cat. No. 564416; Left Plot) or BD Horizon™ BB515 Mouse Anti-Human Siglec-9 (CD329) antibody (Cat. No. 570060/570141; Right Plot). The bivariate pseudocolor density plot showing the correlated expression of Siglec-9 (CD329) [or Ig Isotype control staining] versus side light-scatter (SSC-A) signals was derived from gated events with the forward and side light-scatter characteristics of viable leukocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.

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Product Details
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BD Horizon™
SIGLEC9; siglec-9; CD329
Human (QC Testing)
Mouse BALB/c IgG1, κ
Human Siglec-9-Fc
Flow cytometry (Routinely Tested)
5 µl/test
27180
AB_3685492
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions and unconjugated antibody and free dye were removed. Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze.

Recommended Assay Procedures

   BD® CompBeads can be used as surrogates to assess fluorescence spillover (compensation).  When fluorochrome conjugated antibodies are bound to BD® CompBeads, they have spectral properties very similar to cells.   However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls.  It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.

    For optimal and reproducible results, BD Horizon Brilliant™ Stain Buffer should be used anytime BD Horizon Brilliant™ dyes are used in a multicolor flow cytometry panel.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions. When BD Horizon Brilliant Stain Buffer is used in in the multicolor panel, it should also be used in the corresponding compensation controls for all dyes to achieve the most accurate compensation. For the most accurate compensation, compensation controls created with either cells or beads should be exposed to BD Horizon Brilliant Stain Buffer for the same length of time as the corresponding multicolor panel. More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

For optimal results, it is recommended to perform 2 washes after staining with antibodies. Cells may be prepared, stained with antibodies and washed twice with wash buffer per established protocols for immunofluorescence staining, prior to acquisition on a flow cytometer. Performing fewer than the recommended wash steps may lead to increased spread of the negative population.

Product Notices

  1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  2. This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
  3. An isotype control should be used at the same concentration as the antibody of interest.
  4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
  5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  6. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  7. Alexa Fluor™ is a trademark of Life Technologies Corporation.
  8. For U.S. patents that may apply, see bd.com/patents.

Data Sheets

570060 Rev. 1
Antibody Details
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K8

The K8 monoclonal antibody specifically recognizes Sialic acid-binding Ig-like lectin 9 (Siglec-9) which is also known as CD329. Siglec-9 (CD329) is a single-pass type I transmembrane glycoprotein that is encoded by SIGLEC9 which belongs to the I-type lectins within the Ig superfamily. This lectin is comprised of an N-terminal extracellular region that contains an IgV domain, which binds sialic acid, followed by two Ig-like C2-type domains, a transmembrane region, and an intracellular domain with an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an ITIM-like motif . It is highly expressed on monocytes, neutrophils, and expressed at lower levels on a proportion of NK cells, B cells and T cells. Siglec-9 (CD329) may play roles in the regulation of T cell and NK cell responses and neutrophil apoptosis.

570060 Rev. 1
Format Details
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BB515
The BD Horizon Brilliant™ Blue 515 (BB515) dye is part of the BD Horizon Brilliant™ Blue family of dyes. This dye is a polymer fluorochrome with an excitation maximum (Ex Max) at 490-nm and an emission maximum (Em Max) of 515-nm. Driven by BD innovation, BB515 is designed to be excited by the blue laser (488-nm) and detected using an optical filter centered near 520-nm (e.g., 530/30-nm). BB515 reagents are significantly brighter than equivalent FITC or Alexa Fluor™ 488 reagents with less spillover into the PE detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BB515
490 nm
515 nm
570060 Rev.1
Citations & References
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Development References (5)

  1. Avril T, Floyd H, Lopez F, Vivier E, Crocker PR. The membrane-proximal immunoreceptor tyrosine-based inhibitory motif is critical for the inhibitory signaling mediated by Siglecs-7 and -9, CD33-related Siglecs expressed on human monocytes and NK cells.. J Immunol. 2004; 173(11):6841-9. (Clone-specific: Flow cytometry, Functional assay). View Reference
  2. Carlin AF, Uchiyama S, Chang YC, Lewis AL, Nizet V, Varki A. Molecular mimicry of host sialylated glycans allows a bacterial pathogen to engage neutrophil Siglec-9 and dampen the innate immune response. Blood. 2009; 113(14):3333-3336. (Biology). View Reference
  3. Fraschilla I, Pillai S. Viewing Siglecs through the lens of tumor immunology.. Immunol Rev. 2017; 276(1):178-191. (Biology). View Reference
  4. Nguyen DH, Hurtado-Ziola N, Gagneux P, Varki A. Loss of Siglec expression on T lymphocytes during human evolution.. Proc Natl Acad Sci U S A. 2006; 103(20):7765-70. (Biology). View Reference
  5. Zhang JQ, Nicoll G, Jones C, Crocker PR. Siglec-9, a novel sialic acid binding member of the immunoglobulin superfamily expressed broadly on human blood leukocytes.. J Biol Chem. 2000; 275(29):22121-6. (Immunogen: Flow cytometry). View Reference
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570060 Rev. 1

 

Please refer to Support Documents for Quality Certificates


Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

Please refer to Support Documents for Quality Certificates

 

Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described

 

Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.