Multiparameter flow cytometric analysis of S100A8/A9 (MRP-8/14) expression by human peripheral blood leucocyte populations. Human peripheral blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix cells. After washing, the leucocytes were then permeabilized with and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either Alexa Fluor® 647 Mouse IgG1, κ Isotype Control (Cat. No. 557732; Left Plot) or Alexa Fluor® 647 Mouse Anti-Human S100A8/A9 (MRP-8/14) antibody (Cat. No. 566010; Right Plot). Two-parameter contour plots showing the correlated expression of S100A8/A9 (MRP-8/14) [or Ig Isotype control staining] versus side light-scatter (SSC-A) signals were derived from gated events with the forward and side light-scatter characteristics of viable human peripheral blood leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ X-20 Flow Cytometer System.
Multiparameter flow cytometric analysis of S100A8/A9 (MRP-8/14) expression by human peripheral blood leucocyte populations. Human peripheral blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix cells. After washing, the leucocytes were then permeabilized with and stained in BD Perm/Wash™ Buffer (Cat. No. 554723) with either Alexa Fluor® 647 Mouse IgG1, κ Isotype Control (Cat. No. 557732; Left Plot) or Alexa Fluor® 647 Mouse Anti-Human S100A8/A9 (MRP-8/14) antibody (Cat. No. 566010; Right Plot). Two-parameter contour plots showing the correlated expression of S100A8/A9 (MRP-8/14) [or Ig Isotype control staining] versus side light-scatter (SSC-A) signals were derived from gated events with the forward and side light-scatter characteristics of viable human peripheral blood leucocytes. Flow cytometric analysis was performed using a BD LSRFortessa™ X-20 Flow Cytometer System.
Product Details
BD Pharmingen™
S100A8/S100A9; MRP-8/MRP-14; Calprotectin, CFAG; p8,14
Human (QC Testing)
Mouse BALB/c IgG1, κ
Acute Monocytic Leukemia Cells and Monocytes
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_2732837
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 to Alexa Fluor® 647 under optimum conditions, and unreacted Alexa Fluor® 647 was removed.
Product Notices
- 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).
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
- Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
- Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
- 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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- An isotype control should be used at the same concentration as the antibody of interest.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 mL
Cat No.
554657
Lyse/Fix Buffer 5X RUO
Size
250 mL
Cat No.
558049
Perm/Wash Buffer RUO
Size
100 mL
Cat No.
554723
Alexa Fluor® 647 Mouse IgG1 κ Isotype Control RUO
Size
100 Tests
Cat No.
557732
566010 Rev. 1
Antibody Details
5.5
The 5.5 monoclonal antibody specifically recognizes S100 calcium binding protein A8 (S100A8) and S100A9 (S100A8/A9), which is also known as Migration inhibitory factor-related protein 8 (MRP-8) and MRP-14 (MRP-8/14), Calprotectin, Cystic fibrosis antigen (CF antigen, CFAG) or L1 light and heavy chain. S100A8 and S100A9 are members of the S100 family that regulate myeloid cell functions, inflammation and innate immune responses. S100A8/A9 (MRP-8/14) is expressed in monocytes and neutrophils. Although not generally expressed in tissue macrophages, S100A8/A9 can be detected within some mononuclear cells comprising inflammatory infiltrates. Soluble S100A8/A9 serves as an early serum marker for detecting and monitoring proinflammatory or immunological disease activities.
566010 Rev. 1
Format Details
Alexa Fluor™ 647
Alexa Fluor™ 647 Dye is part of the BD red family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 653-nm and an emission maximum (Em Max) at 669-nm. Alexa Fluor™ 647 is designed to be excited by the Red laser (627-640 nm) and detected using an optical filter centered near 670-nm (e.g., a 660/20 nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 647
Red 627-640 nm
653 nm
669 nm
566010 Rev.1
Citations & References
Development References (6)
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De Ponti A, Wiechert L, Schneller D, et al. A pro-tumorigenic function of S100A8/A9 in carcinogen-induced hepatocellular carcinoma.. Cancer Lett. 2015; 369(2):396-404. (Biology). View Reference
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Hogg N, Allen C, Edgeworth J. Monoclonal antibody 5.5 reacts with p8,14, a myeloid molecule associated with some vascular endothelium.. Eur J Immunol. 1989; 19(6):1053-61. (Immunogen: Immunofluorescence, Immunohistochemistry, Radioimmunoassay). View Reference
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Holzinger D, Frosch M, Kastrup A, et al. The Toll-like receptor 4 agonist MRP8/14 protein complex is a sensitive indicator for disease activity and predicts relapses in systemic-onset juvenile idiopathic arthritis.. Ann Rheum Dis. 2012; 71(6):974-80. (Biology). View Reference
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Linch DC, Allen C, Beverley PC, Bynoe AG, Scott CS, Hogg N. Monoclonal antibodies differentiating between monocytic and nonmonocytic variants of AML.. Blood. 1984; 63(3):566-73. (Immunogen: Immunocytochemistry, Immunofluorescence). View Reference
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Palmer DG, Hogg N, Allen CA, Highton J, Hessian PA. A mononuclear phagocyte subset associated with cell necrosis in rheumatoid nodules: identification with monoclonal antibody 5.5.. Clin Immunol Immunopathol. 1987; 45(1):17-28. (Clone-specific: Immunohistochemistry). View Reference
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Sorg C. The calcium binding proteins MRP8 and MRP14 in acute and chronic inflammation.. Behring Inst Mitt. 1992; (91):126-37. (Biology). View Reference
566010 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.
