Two-color flow cytometric analysis of CD40 expression on mouse and rat splenocytes. Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with PE Rat Anti-Mouse CD5 (Cat. No. 553022/553023) and Alexa Fluor® 647 Hamster Anti-Mouse CD40 (Cat. No. 563638; Left Panel) antibodies. Rat splenic leucocytes were stained with PE Mouse Anti-Rat CD5 (Cat. No. 554851) and Alexa Fluor® 647 Hamster Anti-Mouse CD40 antibodies (Right Panel). The two-color flow cytometric dot plots shows the correlated expression patterns of CD40 versus CD5 for gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Two-color flow cytometric analysis of CD40 expression on mouse and rat splenocytes. Mouse splenic leucocytes were preincubated with Purified Rat Anti-Mouse CD16/CD32 antibody (Mouse BD Fc Block™) (Cat. No. 553141/553142). The cells were then stained with PE Rat Anti-Mouse CD5 (Cat. No. 553022/553023) and Alexa Fluor® 647 Hamster Anti-Mouse CD40 (Cat. No. 563638; Left Panel) antibodies. Rat splenic leucocytes were stained with PE Mouse Anti-Rat CD5 (Cat. No. 554851) and Alexa Fluor® 647 Hamster Anti-Mouse CD40 antibodies (Right Panel). The two-color flow cytometric dot plots shows the correlated expression patterns of CD40 versus CD5 for gated events with the forward and side light-scatter characteristics of viable leucocytes. Flow cytometric analysis was performed using a BD™ LSR II Flow Cytometer System.
Product Details
BD Pharmingen™
Bp50; Tnfrsf5; TNR5; TRAP; CD40L receptor; GP39; HIGM1; IMD3; T-BAM
Mouse (QC Testing), Rat (Tested in Development)
Armenian Hamster IgM, κ
(BALB/c x NZB) F1 Mouse-derived Lymphoma WEHI-231
Flow cytometry (Routinely Tested)
0.2 mg/ml
AB_2738338
Aqueous buffered solution containing protein stabilizer 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
- 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.
- 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.
- Although hamster immunoglobulin isotypes have not been well defined, BD Biosciences Pharmingen has grouped Armenian and Syrian hamster IgG monoclonal antibodies according to their reactivity with a panel of mouse anti-hamster IgG mAbs. A table of the hamster IgG groups, Reactivity of Mouse Anti-Hamster Ig mAbs, may be viewed at http://www.bdbiosciences.com/documents/hamster_chart_11x17.pdf.
- 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® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
- Alexa Fluor® 647 fluorochrome emission is collected at the same instrument settings as for allophycocyanin (APC).
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Species cross-reactivity detected in product development may not have been confirmed on every format and/or application.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
Companion Products
Purified Rat Anti-Mouse CD16/CD32 (Mouse BD Fc Block™) RUO
Size
0.1 mg
Cat No.
553141
Purified Rat Anti-Mouse CD16/CD32 (Mouse BD Fc Block™) RUO
Size
0.5 mg
Cat No.
553142
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Stain Buffer (BSA) RUO
Size
500 mL
Cat No.
554657
PE Rat Anti-Mouse CD5 RUO
Size
0.1 mg
Cat No.
553022
PE Rat Anti-Mouse CD5 RUO
Size
0.2 mg
Cat No.
553023
563638 Rev. 2
Antibody Details
HM40-3
The HM40-3 antibody reacts with CD40, a 40-50-kDa glycoprotein expressed on B lymphocytes and other antigen-presenting cells. The CD40 molecule has a central role in B-cell growth and differentiation. Furthermore, interactions of CD40 with its ligand, CD154, are involved in the initiation and effector stages of cell-mediated immune responses. CD40 may be involved in the triggering of NK cells and NK-T cells. Soluble HM40-3 antibody stimulates splenic and peritoneal B cells to proliferate in vitro. This antibody also induces spleen B cells to express the costimulatory molecules CD80 (B7-1) and CD86 (B7-2). HM40-3 mAb has been demonstrated to inhibit the binding of soluble CD154 (gp39, CD40 Ligand) to soluble CD40 and to cell-surface CD40. This hamster mAb to a mouse leukocyte antigen has been observed to cross-react with similar populations of Lewis, Sprague-Dawley, and LOU16 rat leukocytes.
563638 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 520-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
563638 Rev.2
Citations & References
Development References (15)
-
Akiba H, Oshima H, Takeda K, et al. CD28-independent costimulation of T cells by OX40 ligand and CD70 on activated B cells. J Immunol. 1999; 162(12):7058-7066. (Clone-specific: (Co)-stimulation). View Reference
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Foy TM, Laman JD, Ledbetter JA, Aruffo A, Claassen E, Noelle RJ. gp39-CD40 interactions are essential for germinal center formation and the development of B cell memory. J Exp Med. 1994; 180(1):157-163. (Biology). View Reference
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Grewal IS, Flavell RA. CD40 and CD154 in cell-mediated immunity. Annu Rev Immunol. 1998; 16:111-135. (Biology). View Reference
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Inaba K, Witmer-Pack M, Inaba M, et al. The tissue distribution of the B7-2 costimulator in mice: abundant expression on dendritic cells in situ and during maturation in vitro. J Exp Med. 1994; 180(5):1849-1860. (Immunogen: Flow cytometry). View Reference
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Inaba M, Inaba K, Fukuba Y, et al. Activation of thymic B cells by signals of CD40 molecules plus interleukin-10. Eur J Immunol. 1995; 25(5):1244-1248. (Clone-specific: (Co)-stimulation). View Reference
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Kaneko Y, Hirose S, Abe M, Yagita H, Okumura K, Shirai T. CD40-mediated stimulation of B1 and B2 cells: implication in autoantibody production in murine lupus. Eur J Immunol. 1996; 26(12):3061-3065. (Immunogen: (Co)-stimulation, Flow cytometry, Functional assay). View Reference
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Kashiwada M, Kaneko Y, Yagita H, Okumura K, Takemori T. Activation of mitogen-activated protein kinases via CD40 is distinct from that stimulated by surface IgM on B cells. Eur J Immunol. 1996; 26(7):1451-1458. (Clone-specific: (Co)-stimulation). View Reference
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Kawano T, Cui J, Koezuka Y, et al. CD1d-restricted and TCR-mediated activation of valpha14 NKT cells by glycosylceramides. Science. 1997; 278(5343):1626-1629. (Clone-specific: Blocking). View Reference
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Munder M, Mallo M, Eichmann K, Modolell M. Murine macrophages secrete interferon gamma upon combined stimulation with interleukin (IL)-12 and IL-18: A novel pathway of autocrine macrophage activation. J Exp Med. 1998; 187(12):2103-2108. (Biology). View Reference
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Noelle RJ, Ledbetter JA, Aruffo A. CD40 and its ligand, an essential ligand-receptor pair for thymus-dependent B-cell activation. Immunol Today. 1992; 13(11):431-433. (Biology). View Reference
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Parry SL, Hasbold J, Holman M, Klaus GG. Hypercross-linking surface IgM or IgD receptors on mature B cells induces apoptosis that is reversed by costimulation with IL-4 and anti-CD40. J Immunol. 1994; 152(6):2821-2829. (Biology). View Reference
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Ridge JP, Di Rosa F, Matzinger P. A conditioned dendritic cell can be a temporal bridge between a CD4+ T-helper and a T-killer cell. Nature. 1998; 393(6684):474-478. (Clone-specific: (Co)-stimulation). View Reference
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Tomura M, Yu WG, Ahn HJ, et al. A novel function of Valpha14+CD4+NKT cells: stimulation of IL-12 production by antigen-presenting cells in the innate immune system. J Immunol. 1999; 163(1):93-101. (Biology). View Reference
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Trinite B, Voisine C, Yagita H, Josien R. A subset of cytolytic dendritic cells in rat. J Immunol. 2000; 165(8):4202-4208. (Biology). View Reference
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Turner JG, Rakhmilevich AL, Burdelya L, et al. Anti-CD40 antibody induces antitumor and antimetastatic effects: the role of NK cells. J Immunol. 2001; 166(1):89-94. (Biology). View Reference
563638 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.
