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BUV737 Rat Anti-Mouse CD14
BUV737 Rat Anti-Mouse CD14
Flow cytometric analysis of CD14 expression on mouse J774A.1 cells. Cells from the J774A.1 (Mouse macrophage, ATCC TIB-67) cell line were stained with either BD Horizon™ BUV737 Rat IgG1, κ Isotype Control (Cat. No. 564690; dashed line histogram) or BD Horizon BUV737 Rat Anti-Mouse CD14 antibody (Cat. No. 565309; solid line histogram).  The fluorescence histogram showing CD14 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable J774A cells. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Flow cytometric analysis of CD14 expression on mouse J774A.1 cells. Cells from the J774A.1 (Mouse macrophage, ATCC TIB-67) cell line were stained with either BD Horizon™ BUV737 Rat IgG1, κ Isotype Control (Cat. No. 564690; dashed line histogram) or BD Horizon BUV737 Rat Anti-Mouse CD14 antibody (Cat. No. 565309; solid line histogram).  The fluorescence histogram showing CD14 expression (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable J774A cells. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Product Details
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BD Horizon™
Cd14; CD14 antigen; Myeloid cell-specific leucine-rich glycoprotein
Mouse (QC Testing)
Rat LOU, also known as Louvain, LOU/C, LOU/M IgG1, κ
Recombinant Mouse CD14
Flow cytometry (Routinely Tested)
0.2 mg/ml
12475
AB_2739175
Aqueous buffered solution containing ≤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 BUV737 under optimum conditions, and unconjugated antibody and free BD Horizon BUV737 were removed.

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 CompBead 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 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) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

Note:  When using high concentrations of antibody, background binding of this dye to erythroid cell subsets (mature erythrocytes and precursors) has been observed.  For researchers studying these cell populations, or in cases where light scatter gating does not adequately exclude these cells from the analysis, this background may be an important factor to consider when selecting reagents for panel(s).

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. 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.
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  6. 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.
  7. BD Horizon Brilliant Ultraviolet 737 is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,227,187; 8,575,303; 8,354,239.
  8. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
565309 Rev. 2
Antibody Details
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rmC5-3

The rmC5-3 monoclonal antibody specifically binds to residues 308-322 of the hydrophilic region of mouse CD14. CD14 is a 53-55 kDa glycophosphatidylinositol (GPI)-linked glycoprotein belonging to the leucine-rich glycoprotein repeat superfamily of cell-surface proteins. It is a receptor for the complex of lipopolysaccharide (LPS or endotoxin, from gram-negative bacteria) with LPS-binding protein (LBP, a plasma protein). CD14 serves as a receptor for LPS that can play a role in the cellular production of proinflammatory cytokines such as IL-1 and TNF. CD14 can be involved in the development of endotoxic shock and LPS-stimulated bone resorption, and promotes, possibly indirectly, bacterial dissemination. Flow cytometric analysis demonstrates that rmC5-3 antibody stains J774A.1 (mouse macrophage line), WEHI-265.1 (mouse monocytic line), peritoneal resident macrophages, Kupffer cells, and cultured bone marrow-derived macrophages and dendritic cells, but not unstimulated splenic macrophages, dendritic cells, neutrophils, or blood monocytes. This staining pattern is similar to that of the alternate anti-mouse CD14 mAb 4C1/CD14, which recognizes a different CD14 epitope, and differs from that of the human, where CD14 expression is characteristic of circulating monocytes and neutrophils. Therefore, data suggests that CD14 expression by leukocyte populations may differ in mice and humans. Peritoneal cells from naive mice, 3-day thioglycollate-elicited peritoneal exudate, as well as 4-hour LPS-activated peritoneal cells, contain a population of Mac-1 (CD11b)-high cells which double-stain with rmC5-3 antibody. Levels of CD14 expression on Kupffer cells and bone marrow-derived macrophages and dendritic cells of LPS-sensitive mice are increased by in vivo and in vitro LPS treatments, an effect which may be mediated by TNF. Preliminary evidence suggests that CD14 may be up-regulated on mouse blood neutrophils. In agreement with the observations that CD14 is shed from activated human and mouse monocytes, rmC5-3 mAb detects soluble CD14 in the serum of LPS-treated mice in a time-dependent manner.

The antibody was conjugated to BD Horizon BUV737 which is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This dye is a tandem fluorochrome with an Ex Max near 350 nm and an Em Max near 737 nm. BD Horizon Brilliant BUV737 can be excited by the ultraviolet laser (355 nm) and detected with a 740/35 nm filter.  Due to the excitation of the acceptor dye by the red laser line, there may be significant spillover into red laser detectors with filters in the 700-720 nm range.

565309 Rev. 2
Format Details
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BUV737
The BD Horizon Brilliant™ Ultraviolet 737 (BUV737) Dye is part of the BD Horizon Brilliant™ Ultraviolet family of dyes. This tandem fluorochrome is comprised of a BUV395 donor with an excitation maximum (Ex Max) of 350-nm and an acceptor dye with an emission maximum (Em Max) at 735-nm. BUV737, driven by BD innovation, is designed to be excited by the ultraviolet laser (355-nm) and detected using an optical filter centered near 740-nm (e.g., 740/35 bandpass filter). The acceptor dye can be excited by the Red (628–640nm) laser resulting in cross-laser excitation and fluorescence spillover. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BUV737
Ultraviolet 355 nm
350 nm
735 nm
565309 Rev.2
Citations & References
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View product citations for antibody "565309" on CiteAb

Development References (13)

  1. Fearns C, Kravchenko VV, Ulevitch RJ, Loskutoff DJ. Murine CD14 gene expression in vivo: extramyeloid synthesis and regulation by lipopolysaccharide. J Exp Med. 1995; 181(3):857-866. (Biology). View Reference
  2. Fearns C, Loskutoff DJ. Role of tumor necrosis factor alpha in induction of murine CD14 gene expression by lipopolysaccharide. Infect Immun. 1997; 65(11):4822-4831. (Biology). View Reference
  3. Ferrero E, Hsieh CL, Francke U, Goyert SM. CD14 is a member of the family of leucine-rich proteins and is encoded by a gene syntenic with multiple receptor genes. J Immunol. 1990; 145(1):331-336. (Biology). View Reference
  4. Haziot A, Ferrero E, Kontgen F, et al. Resistance to endotoxin shock and reduced dissemination of gram-negative bacteria in CD14-deficient mice. Immunity. 1996; 4(4):407-414. (Clone-specific: Flow cytometry). View Reference
  5. Mahnke K, Becher P, Ricciardi-Castagnoli P, Luger TA, Schawrz T Grabbe S. CD14 is expressed by subsets of murine dendritic cells and upregulated by lipopolysaccharide. In: Ricciardi-Castagnoli P, ed. Dendritic Cells in Fundamental and Clinical Immunology. New York: Plenum Press; 1997:145-159.
  6. Matsuura K, Ishida T, Setoguchi M, Higuchi Y, Akizuki S, Yamamoto S. Upregulation of mouse CD14 expression in Kupffer cells by lipopolysaccharide. J Exp Med. 1994; 179(5):1671-1676. (Immunogen: ELISA, Flow cytometry, Immunohistochemistry, Western blot). View Reference
  7. Miyata Y, Takeda H, Kitano S, Hanazawa S. Porphyromonas gingivalis lipopolysaccharide-stimulated bone resorption via CD14 is inhibited by broad-spectrum antibiotics. Infect Immun. 1997; 65(9):3513-3519. (Clone-specific: Immunohistochemistry). View Reference
  8. Nasu N, Yoshida S, Akizuki S, Higuchi Y, Setoguchi M, Yamamoto S. Molecular and physiological properties of murine CD14. Int Immunol. 1991; 3(2):205-213. (Biology). View Reference
  9. Pulendran B, Lingappa J, Kennedy MK, et al. Developmental pathways of dendritic cells in vivo: distinct function, phenotype, and localization of dendritic cell subsets in FLT3 ligand-treated mice. J Immunol. 1997; 159(5):2222-2231. (Clone-specific: Flow cytometry). View Reference
  10. Stewart CC. Methods for studying the ontogeny of monnuclear phagocytes. In: Weir DM, Herzenberg LA, Blackwell C, ed. Weir's Handbook of Experiemental Immunology. Blackwell Science Publications; 1986:44.1-44.17.
  11. Takakuwa T, Knopf HP, Sing A, Carsetti R, Galanos C, Freudenberg MA. Induction of CD14 expression in Lpsn, Lpsd and tumor necrosis factor receptor-deficient mice. Eur J Immunol. 1996; 26(11):2686-2692. (Clone-specific: Flow cytometry). View Reference
  12. Takamatsu S, Nakashima I, Nakano K. Modulation of endotoxin-induced histamine synthesis by cytokines in mouse bone marrow-derived macrophages. J Immunol. 1996; 156(2):778-785. (Clone-specific: Flow cytometry). View Reference
  13. Takezawa R, Watanabe Y, Akaike T. Direct evidence of macrophage differentiation from bone marrow cells in the liver: a possible origin of Kupffer cells. J Biochem (Tokyo). 1995; 118(6):1175-1183. (Biology). View Reference
View All (13) View Less
565309 Rev. 2

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