BD Horizon™ RB744 Rat Anti-Mouse Siglec-H
Clone 440c (RUO)
Multiparameter flow cytometric analysis of Siglec-H expression on Mouse leukocytes. 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 leukocytes (Panel B) were stained with PE Rat Anti-Mouse CD45R/B220 (Cat. No. 553090) and with either BD Horizon™ RB744 Rat IgG2b, κ Isotype Control (Cat. No. 570521; Top Plots) or BD Horizon™ RB744 Rat Anti-Mouse Siglec-H antibody (Cat. No. 570576/570664; Bottom Plots) at 0.25 µg/test. DAPI Solution (Cat. No. 564907) was added to cells right before analysis. Bivariate pseudocolor density plots showing the correlated expression of Siglec-H (or Ig Isotype control staining) versus CD45R/B220 were derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leukocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software. Data shown on this Technical Data Sheet are not lot specific.
Multiparameter flow cytometric analysis of Siglec-H expression on Mouse leukocytes. 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 leukocytes (Panel B) were stained with PE Rat Anti-Mouse CD45R/B220 (Cat. No. 553090) and with either BD Horizon™ RB744 Rat IgG2b, κ Isotype Control (Cat. No. 570521; Top Plots) or BD Horizon™ RB744 Rat Anti-Mouse Siglec-H antibody (Cat. No. 570576/570664; Bottom Plots) at 0.25 µg/test. DAPI Solution (Cat. No. 564907) was added to cells right before analysis. Bivariate pseudocolor density plots showing the correlated expression of Siglec-H (or Ig Isotype control staining) versus CD45R/B220 were derived from gated events with the forward and side light-scatter characteristics of viable (DAPI-negative) leukocytes. Flow cytometry and data analysis were performed using a BD FACSymphony™ A5 SE Cell Analyzer System and FlowJo™ Software. Data shown on this Technical Data Sheet are not lot specific.
Product Details
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)
0.2 mg/ml
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 unreacted dye was 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.
Product Notices
- When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- An isotype control should be used at the same concentration as the antibody of interest.
- Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
- For U.S. patents that may apply, see bd.com/patents.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 mL
Cat No.
554657
RB744 Rat IgG2b, κ Isotype Control RUO
Size
0.1 mg
Cat No.
570521
Purified Rat Anti-Mouse CD16/CD32 (Mouse BD Fc Block™) RUO
Size
0.5 mg
Cat No.
553142
DAPI Solution RUO
Size
1 mg
Cat No.
564907
Lysing Buffer RUO
Size
100 mL
Cat No.
555899
570576 Rev. 1
Antibody Details
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.
570576 Rev. 1
Format Details
RB744
The BD Horizon RealBlue™ 744 (RB744) Dye is part of the BD® family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 498-nm and an emission maximum (Em Max) at 746-nm as measured using an antibody-dye conjugate. Driven by BD® innovation, RB744 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with minimal excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), we recommend using an optical filter centered near 750-nm (e.g., a 750/60-nm bandpass filter).
RB744
Blue 488 nm
498 nm
746 nm
570576 Rev.1
Citations & References
View product citations for antibody "570576" on CiteAb
Development References (8)
-
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
-
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
-
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
-
Schmitt H, Sell S, Koch J, et al. Siglec-H protects from virus-triggered severe systemic autoimmunity.. J Exp Med. 2016; 213(8):1627-44. (Biology). View Reference
-
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):955-66. (Clone-specific: Flow cytometry). View Reference
-
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):4409-16. (Clone-specific: Flow cytometry, Fluorescence activated cell sorting). View Reference
-
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):2367-74. (Clone-specific: Immunohistochemistry). View Reference
-
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):3600-8. (Biology). View Reference
570576 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.
For Research Use Only. Not for use in diagnostic or therapeutic procedures.