Multiparameter flow cytometric analysis of Myeloperoxidase expression in Human peripheral blood leucocyte populations. Whole blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix leucocytes. The fixed leucocytes were washed and then permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The permeabilized cells were stained in BD Perm/Wash™ Buffer with either Alexa Fluor™ 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; Left Plot) or Alexa Fluor™ 647 Mouse Anti-Human Myeloperoxidase antibody (Cat. No. 568914/568915; Right Plot). The bivariate pseudocolor density plot showing the correlated expression of Myeloperoxidase (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 intact leucocyte populations. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.
Multiparameter flow cytometric analysis of Myeloperoxidase expression in Human peripheral blood leucocyte populations. Whole blood was treated with BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049) to lyse erythrocytes and fix leucocytes. The fixed leucocytes were washed and then permeabilized with BD Perm/Wash™ Buffer (Cat. No. 554723). The permeabilized cells were stained in BD Perm/Wash™ Buffer with either Alexa Fluor™ 647 Mouse IgG1 κ Isotype Control (Cat. No. 557732; Left Plot) or Alexa Fluor™ 647 Mouse Anti-Human Myeloperoxidase antibody (Cat. No. 568914/568915; Right Plot). The bivariate pseudocolor density plot showing the correlated expression of Myeloperoxidase (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 intact leucocyte populations. Flow cytometry and data analysis were performed using a BD LSRFortessa™ X-20 Cell Analyzer System and FlowJo™ software.
Product Details
BD Pharmingen™
Myeloperoxidase; MPO
Human (QC Testing)
Mouse BALB/c IgG1, κ
Purified Human Myeloperoxidase
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
4353
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 the dye under optimum conditions and unreacted dye was 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 cell and BD® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
Product Notices
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
- 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.
- 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).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- An isotype control should be used at the same concentration as the antibody of interest.
- This product is provided under an intellectual property license between Life Technologies Corporation and BD Businesses. The purchase of this product conveys to the buyer the non-transferable right to use the purchased amount of the product and components of the product in research conducted by the buyer (whether the buyer is an academic or for-profit entity). The buyer cannot sell or otherwise transfer (a) this product (b) its components or (c) materials made using this product or its components to a third party or otherwise use this product or its components or materials made using this product or its components for Commercial Purposes. Commercial Purposes means any activity by a party for consideration and may include, but is not limited to: (1) use of the product or its components in manufacturing; (2) use of the product or its components to provide a service, information, or data; (3) use of the product or its components for therapeutic, diagnostic or prophylactic purposes; or (4) resale of the product or its components, whether or not such product or its components are resold for use in research. For information on purchasing a license to this product for any other use, contact Life Technologies Corporation, Cell Analysis Business Unit Business Development, 29851 Willow Creek Road, Eugene, OR 97402, USA, Tel: (541) 465-8300. Fax: (541) 335-0504.
- Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
- Alexa Fluor™ is a trademark of Life Technologies Corporation.
- 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
Alexa Fluor® 647 Mouse IgG1 κ Isotype Control RUO
Size
100 Tests
Cat No.
557732
Lyse/Fix Buffer 5X RUO
Size
250 mL
Cat No.
558049
Perm/Wash Buffer RUO
Size
100 mL
Cat No.
554723
568914 Rev. 2
Antibody Details
5B8
The 5B8 monoclonal antibody specifically recognizes Myeloperoxidase (MPO). Enzymatically active Myeloperoxidase exists as a heterotetrameric glycoprotein comprised of two disulfide-linked heterodimers with each having a heavy (~55 kDa) and light (~15 kDa) subunit associated with a heme-like prosthetic group. Myeloperoxidase is stored in primary azurophilic granules of neutrophils and is rapidly released into the phagosome and extracellular space during inflammatory responses. It is also present in monocytes and in myeloblasts from some cases of acute myeloid leukemia. Myeloperoxidase catalyzes the generation of hypochlorous acid, a powerful oxidant and potent microbicidal agent. It plays a major role in the bactericidal activity of neutrophils and monocytes.
568914 Rev. 2
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
568914 Rev.2
Citations & References
View product citations for antibody "568914" on CiteAb
Development References (7)
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Audrain MA, Baranger TA, Moguilevski N, et al. Anti-native and recombinant myeloperoxidase monoclonals and human autoantibodies. Clin Exp Immunol. 1997; 107(1):127-134. (Immunogen: Blocking, ELISA, Western blot). View Reference
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He Z, Allers C, Sugimoto C, et al. Rapid Turnover and High Production Rate of Myeloid Cells in Adult Rhesus Macaques with Compensations during Aging.. J Immunol. 2018; 200(12):4059-4067. (Clone-specific: Flow cytometry). View Reference
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Knapp W, Strobl H, Majdic O. Flow cytometric analysis of cell-surface and intracellular antigens in leukemia diagnosis. Cytometry. 1994; 18(4):187-198. (Biology). View Reference
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Raskovalova T, Berger MG, Jacob MC, et al. Flow cytometric analysis of neutrophil myeloperoxidase expression in peripheral blood for ruling out myelodysplastic syndromes: a diagnostic accuracy study.. Haematologica. 2019; 104(12):2382-2390. (Clone-specific: Flow cytometry). View Reference
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Taylor KL, Pohl J, Kinkade JM. Unique autolytic cleavage of human myeloperoxidase. Implications for the involvement of active site MET409.. J Biol Chem. 1992; 267(35):25282-8. (Biology). View Reference
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Tobler A, Miller CW, Johnson KR, Selsted ME, Rovera G, Koeffler HP. Regulation of gene expression of myeloperoxidase during myeloid differentiation. J Cellular Physiol. 1988; 136:215-225. (Biology).
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van de Vyver M, Engelbrecht L, Smith C, Myburgh KH. Neutrophil and monocyte responses to downhill running: Intracellular contents of MPO, IL-6, IL-10, pstat3, and SOCS3.. Scand J Med Sci Sports. 2016; 26(6):638-47. (Clone-specific: Flow cytometry). View Reference
568914 Rev. 2
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.