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BV421 Rat Anti-Mouse Siglec-H
BV421 Rat Anti-Mouse Siglec-H
Analyses of Siglec-H expression.        Panels A and B. Flow cytometric analysis of Siglec-H expression on mouse leucocytes. SJL mouse splenocytes (Panel A; preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody/Mouse BD Fc Block™, Cat. No. 553141/553142) and BALB/c thymic leucocytes (Panel B) were stained with Alexa Fluor® 647 Rat Anti-Mouse CD45R/B220 (Cat. No. 557683) and either BD Horizon™ BV421 Rat IgG2b, κ (Cat. No. 562603; Top Plots) or BD Horizon BV421 Rat Anti-Mouse Siglec-H antibody (Cat. No. 566581; Bottom Plots) at 0.125 µg/test. Flow cytometric contour plots showing the correlated expression of Siglec-H (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa Flow Cytometry System. Data shown on this Technical Data Sheet are not lot specific.        Panel C. Immunohistofluorescent staining of Siglec-H in mouse spleen and thymus. BALB/c mouse frozen spleen (Top Image) and thymus (Bottom Image) sections were fixed with cold acetone, washed, and then stained with BD Horizon™ BV421 Rat Anti-Mouse Siglec -H (pseudocolored red) and FITC Hamster Anti-Mouse CD3e (Cat. No.561827, pseudocolored green) antibodies, and in addition for spleen, Alexa 647® Rat Anti-Mouse CD19 antibody (Cat. No. 557684, pseudocolored blue). The images were generated using a Molecular Devices Standard Epifluorescence microscope. Original magnification, 20x.
Analyses of Siglec-H expression.        Panels A and B. Flow cytometric analysis of Siglec-H expression on mouse leucocytes. SJL mouse splenocytes (Panel A; preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody/Mouse BD Fc Block™, Cat. No. 553141/553142) and BALB/c thymic leucocytes (Panel B) were stained with Alexa Fluor® 647 Rat Anti-Mouse CD45R/B220 (Cat. No. 557683) and either BD Horizon™ BV421 Rat IgG2b, κ (Cat. No. 562603; Top Plots) or BD Horizon BV421 Rat Anti-Mouse Siglec-H antibody (Cat. No. 566581; Bottom Plots) at 0.125 µg/test. Flow cytometric contour plots showing the correlated expression of Siglec-H (or Ig Isotype control staining) were derived from gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa Flow Cytometry System. Data shown on this Technical Data Sheet are not lot specific.        Panel C. Immunohistofluorescent staining of Siglec-H in mouse spleen and thymus. BALB/c mouse frozen spleen (Top Image) and thymus (Bottom Image) sections were fixed with cold acetone, washed, and then stained with BD Horizon™ BV421 Rat Anti-Mouse Siglec -H (pseudocolored red) and FITC Hamster Anti-Mouse CD3e (Cat. No.561827, pseudocolored green) antibodies, and in addition for spleen, Alexa 647® Rat Anti-Mouse CD19 antibody (Cat. No. 557684, pseudocolored blue). The images were generated using a Molecular Devices Standard Epifluorescence microscope. Original magnification, 20x.
Product Details
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BD Horizon™
Siglec H; Siglech; Sialic acid binding Ig-like lectin H; SIGLEC
Mouse (QC Testing)
Rat IgG2b, κ
Mouse Bone Marrow IPC
Flow cytometry (Routinely Tested), Immunofluorescence (Tested During Development)
0.2 mg/ml
233274
AB_2739747
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated with BD Horizon BV421 under optimum conditions, and unconjugated antibody and free BD Horizon BV421 were removed.

Recommended Assay Procedures

For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes are used in the same experiment.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions.  More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349).

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  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. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  7. BD Horizon Brilliant Violet 421 is covered by one or more of the following US patents: 8,158,444; 8,362,193; 8,575,303; 8,354,239.
  8. BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
  9. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
566581 Rev. 1
Antibody Details
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440c

The 440c monoclonal antibody specifically recognizes Siglec-H, a type I transmembrane glycoprotein that is encoded by Siglech (Sialic acid binding Ig-like lectin H). Siglec-H contains two immunoglobulin domains in its extracellular region and a cytoplasmic domain that lacks tyrosine-based signaling motifs unlike other CD33-related Siglec-like molecules. Siglec-H is expressed on progenitors of plasmacytoid dendritic cells (pDCs) and depends on DAP12 for cell surface expression and intracellular signaling function. In addition to its expression on pDC, Siglec-H may also be expressed by splenic marginal zone macrophages, medullary macrophages in lymph nodes, and microglia. Siglec-H may regulate the production of type I interferons (IFN-I) by pDCs and other cell types. There is no clear human ortholog for mouse Siglec-H.

        The antibody was conjugated to BD Horizon BV421 which is part of the BD Horizon Brilliant™ Violet family of dyes.  With an Ex Max of 407-nm and Em Max at 421-nm, BD Horizon BV421  can be excited by the violet laser and detected in the standard Pacific Blue™ filter set (eg, 450/50-nm filter). BD Horizon BV421  conjugates are very bright, often exhibiting a 10 fold improvement in brightness compared to Pacific Blue conjugates.

566581 Rev. 1
Format Details
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BV421
The BD Horizon Brilliant Violet™ 421 (BV421) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This polymer-technology based dye has an excitation maximum (Ex Max) of 407-nm and an emission maximum (Em Max) at 423-nm. Driven by BD innovation, BV421 is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 420-nm (e.g., a 431/28-nm or 450/50-nm bandpass filter). BV421 is an ideal alternative for V450 as it is approximately ten times brighter with less spillover into the BV510/V500 detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
altImg
BV421
Violet 405 nm
407 nm
423 nm
566581 Rev.1
Citations & References
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Development References (8)

  1. Blasius A, Vermi W, Krug A, Facchetti F, Cella M, Colonna M. A cell-surface molecule selectively expressed on murine natural interferon-producing cells that blocks secretion of interferon-alpha.. Blood. 2004; 103(11):4201-6. (Immunogen: Flow cytometry, Fluorescence activated cell sorting, Functional assay, Immunohistochemistry, Inhibition, In vivo exacerbation). View Reference
  2. Blasius AL, Cella M, Maldonado J, Takai T, Colonna M. Siglec-H is an IPC-specific receptor that modulates type I IFN secretion through DAP12.. Blood. 2006; 107(6):2474-6. (Clone-specific: Flow cytometry). View Reference
  3. Blasius AL, Giurisato E, Cella M, Schreiber RD, Shaw AS, Colonna M. Bone marrow stromal cell antigen 2 is a specific marker of type I IFN-producing cells in the naive mouse, but a promiscuous cell surface antigen following IFN stimulation.. J Immunol. 2006; 177(5):3260-5. (Clone-specific: Flow cytometry). View Reference
  4. Schmitt H, Sell S, Koch J, et al. Siglec-H protects from virus-triggered severe systemic autoimmunity. J Exp Med. 2016; 213(8):1627-1644. (Biology). View Reference
  5. Swiecki M, Gilfillan S, Vermi W, Wang Y, Colonna M. Plasmacytoid dendritic cell ablation impacts early interferon responses and antiviral NK and CD8(+) T cell accrual. Immunity. 2010; 33(6):955-966. (Clone-specific: Flow cytometry). View Reference
  6. Swiecki M, Wang Y, Riboldi E, et al. Cell depletion in mice that express diphtheria toxin receptor under the control of SiglecH encompasses more than plasmacytoid dendritic cells. J Immunol. 2014; 192(9):4409-4416. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
  7. Swiecki M, Wang Y, Vermi W, Gilfillan S, Schreiber RD, Colonna M. Type I interferon negatively controls plasmacytoid dendritic cell numbers in vivo. J Exp Med. 2011; 208(12):2367-2374. (Clone-specific: Immunohistochemistry). View Reference
  8. Zhang J, Raper A, Sugita N, et al. Characterization of Siglec-H as a novel endocytic receptor expressed on murine plasmacytoid dendritic cell precursors. Blood. 2006; 107(9):3600-3608. (Biology). View Reference
View All (8) View Less
566581 Rev. 1

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