Multiparameter flow cytometric analysis of CD40 expression on Human peripheral blood leucocyte populations. Platelet-depleted human whole blood was stained with either Alexa Fluor™ 488 Mouse IgG1, κ Isotype Control (Cat. No. 565572; Left Plot) or Alexa Fluor™ 488 Mouse Anti-Human CD40 antibody (Cat. No. 568964/568965; Right Plot). Erythrocytes were lysed with BD FACS Lysing™ Solution (Cat. No. 349202). The bivariate pseudocolor density plot showing the correlated expression of CD40 (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.
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Multiparameter flow cytometric analysis of CD40 expression on Human peripheral blood leucocyte populations. Platelet-depleted human whole blood was stained with either Alexa Fluor™ 488 Mouse IgG1, κ Isotype Control (Cat. No. 565572; Left Plot) or Alexa Fluor™ 488 Mouse Anti-Human CD40 antibody (Cat. No. 568964/568965; Right Plot). Erythrocytes were lysed with BD FACS Lysing™ Solution (Cat. No. 349202). The bivariate pseudocolor density plot showing the correlated expression of CD40 (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™
CD40; TNFRSF5; Bp50
Human (QC Testing), Rhesus,Cynomolgus,Baboon (Reported)
Mouse IgG1, κ
Human CD40
Flow cytometry (Routinely Tested)
5 µl
V B CD40.5; VIII 377
958
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 cells 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.
- 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).
- An isotype control should be used at the same concentration as the antibody of interest.
- Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
- 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.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- 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.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 mL
Cat No.
554657
Alexa Fluor® 488 Mouse IgG1 κ Isotype Control RUO
Size
50 µg
Cat No.
565572
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Lysing Solution 10X Concentrate IVD
Size
100 mL
Cat No.
349202
Lysing Buffer RUO
Size
100 mL
Cat No.
555899
568965 Rev. 1
Antibody Details
HB14
This HB14 monoclonal antibody specifically recognizes CD40 which is also known as Tumor necrosis factor receptor superfamily member 5 (TNFRSF5). CD40 is an ~45-48 kDa single-pass type I transmembrane glycoprotein. CD40 is expressed on B lymphocytes, monocytes, macrophages, dendritic cells, endothelial cells, fibroblasts, keratinocytes, and CD34+ hematopoietic progenitor cells. CD40 serves as a receptor for CD154 (CD40 Ligand or CD40L). CD40:CD154 interactions costimulate B cell growth, differentiation, and immunoglobulin isotype switching in antibody responses to T-dependent antigens as well as the generation of memory B cells and cytokine production by monocytes and dendritic cells. The HB14 antibody reportedly crossreacts with CD40 expressed by several non-human primates including Rhesus and Cynomolgus Macaques as well as Olive Baboons.
568965 Rev. 1
Format Details
Alexa Fluor™ 488
Alexa Fluor™ 488 Dye is part of the BD blue family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 494-nm and an emission maximum (Em Max) at 517-nm. Alexa Fluor™ 488 is designed to be excited by the Blue laser (488 nm) and detected using an optical filter centered near 520-nm (e.g., a 530/30-nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 488
Blue 488 nm
494 nm
517 nm
568965 Rev.1
Citations & References
View product citations for antibody "568965" on CiteAb
Development References (12)
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Barclay NA, Brown MH, Birkeland ML, et al, ed. The Leukocyte Antigen FactsBook. San Diego, CA: Academic Press; 1997.
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Bjornson-Hooper ZB, Fragiadakis GK, Spitzer MH, et al. A Comprehensive Atlas of Immunological Differences Between Humans, Mice, and Non-Human Primates.. Front Immunol. 2022; 13:867015. (Clone-specific: Flow cytometry). View Reference
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Clark EA, Ledbetter JA. Activation of human B cells mediated through two distinct cell surface differentiation antigens, Bp35 and Bp50. Proc Natl Acad Sci U S A. 1986; 83(12):4494-4498. (Biology). View Reference
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Freedman JE. CD40-CD40L and platelet function: beyond hemostasis.. Circ Res. 2003; 92(9):944-6. (Biology). View Reference
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Galy AH, Spits H. CD40 is functionally expressed on human thymic epithelial cells. J Immunol. 1992; 149(3):775-782. (Biology). View Reference
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Hu HM, O'Rourke K, Boguski MS, Dixit VM. A novel RING finger protein interacts with the cytoplasmic domain of CD40.. J Biol Chem. 1994; 269(48):30069-72. (Clone-specific: Immunoprecipitation). View Reference
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Inwald DP, McDowall A, Peters MJ, Callard RE, Klein NJ. CD40 is constitutively expressed on platelets and provides a novel mechanism for platelet activation.. Circ Res. 2003; 92(9):1041-8. (Biology). View Reference
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Itoh N, Yonehara S, Ishii A, et al. The polypeptide encoded by the cDNA for human cell surface antigen Fas can mediate apoptosis. Cell. 1991; 66(2):233-243. (Biology). View Reference
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Kansas GS, Tedder TF. Transmembrane signals generated through MHC class II, CD19, CD20, CD39, and CD40 antigens induce LFA-1-dependent and independent adhesion in human B cells through a tyrosine kinase-dependent pathway. J Immunol. 1991; 147(12):4094-4102. (Immunogen: Functional assay). View Reference
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Mitchell K, Barreyro L, Todorova TI, et al. IL1RAP potentiates multiple oncogenic signaling pathways in AML.. J Exp Med. 2018; 215(6):1709-1727. (Clone-specific: Flow cytometry). View Reference
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Pound JD, Challa A, Holder MJ, et al. Minimal cross-linking and epitope requirements for CD40-dependent suppression of apoptosis contrast with those for promotion of the cell cycle and homotypic adhesions in human B cells.. Int Immunol. 1999; 11(1):11-20. (Clone-specific: Functional assay). View Reference
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Schlossman SF. Stuart F. Schlossman .. et al., ed. Leucocyte typing V : white cell differentiation antigens : proceedings of the fifth international workshop and conference held in Boston, USA, 3-7 November, 1993. Oxford: Oxford University Press; 1995.
568965 Rev. 1
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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.