Invitrogen™ CD40 Monoclonal Antibody (5C3), Super Bright™ 702, eBioscience™

Catalog No.	Quantity
67-040-942	100 Tests

Catalog No. 67-040-942 Supplier Invitrogen™ Supplier No. 67040942

Description

Mouse Monoclonal Antibody

Description: The 5C3 monoclonal antibody reacts with human CD40, a 45-50 kDa type I transmembrane glycoprotein. CD40 is a member of the TNFR family and is expressed by B lymphocytes, follicular dendritic cells, thymic epithelium, and a subset of peripheral T cells. CD40 regulates B cell development and maturation by inducing Ig isotype-switching and in combination with other signals such as IL-4, protects B cells from surface Ig-induced apoptosis and promotes proliferation. Interaction of CD40 with CD154 (gp39), its ligand on T cells, is important in T-B cell crosstalk and plays a role in costimulation and immune regulation. 5C3 is reported to be used for activation of B cells in in vitro functional assays. Applications Reported: This 5C3 antibody has been reported for use in flow cytometric analysis. Applications Tested: This 5C3 antibody has been tested by flow cytometric analysis of normal human peripheral blood cells. This may be used at less than or equal to 5 μL (0.125) μg per test. A test is defined as the amount (μg) of antibody that will stain a cell sample in a final volume of 100 μL. Cell number should be determined empirically but can range from 10^5 to 10^8 cells/test. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. Super Bright 702 is a tandem dye that can be excited with the violet laser line (405 nm) and emits at 702 nm. We recommend using a 710/50 bandpass filter. Please make sure that...

CD40 is a single-chain glycoprotein and a member of the tumor necrosis factor receptor (TNFR) family, exhibiting significant homology to the Hodgkin's disease-associated antigen, CD30. It is expressed by B lymphocytes, follicular dendritic cells, thymic epithelium, and a subset of peripheral T cells, as well as some epithelial cells, carcinomas, and lymphoid dendritic cells. Notably, CD40 is present on all B cells except plasma cells. CD40 plays a crucial role in regulating B cell development and maturation, inducing immunoglobulin isotype-switching, and protecting B cells from surface Ig-induced apoptosis when combined with other signals such as IL-4. It promotes proliferation and is essential for T cell-dependent immunoglobulin class switching, memory B cell development, and germinal center formation. The interaction between CD40 and its ligand CD154 (gp39) on T cells is vital for T-B cell crosstalk, costimulation, and immune regulation. Adaptor protein TNFR2 interacts with CD40, mediating signal transduction, while the AT-hook transcription factor AKNA is reported to regulate CD40 expression, which may be important for homotypic cell interactions. Additionally, the interaction between CD40 and its ligand is necessary for amyloid-beta-induced microglial activation, suggesting a role in the early pathogenesis of Alzheimer's disease. Two alternatively spliced transcript variants encoding distinct isoforms of CD40 have been identified. Diseases associated with CD40 dysfunction include Type 3 Hyper-IgM immunodeficiency and CD40 ligand deficiency, highlighting its importance in immune and inflammatory responses.

Specifications

• Antigen: CD40
• Applications: Flow Cytometry
• Classification: Monoclonal
• Clone: 5C3
• Concentration: 5 μL/Test
• Conjugate: Super Bright 702
• Formulation: PBS with BSA and 0.09% sodium azide; pH 7.2
• Gene: CD40
• Gene Accession No.: P25942
• Gene Alias: AI326936; B cell surface antigen CD40; B cell-associated molecule; B-cell surface antigen CD40; Bp50; CD antigen CD40; Cd40; CD40 antigen; CD40 antigen (TNF receptor superfamily member 5); CD40 antigen, TNF receptor superfamily member 5; CD40 molecule; CD40 molecule, TNF receptor superfamily member 5; CD40L receptor; CDw40; GP39; HIGM1; I79_006806; IGM; IMD3; Immunoglobulin M; ImmunoglobulinM; membrane protein CD40; MGC9013; p50; receptor for ligand CD154; sCD40; soluble CD40; T-BAM; T-cell differentiation antigen; Tnfrsf5; TRAP; tumor necrosis factor receptor superfamily member 5; tumor necrosis factor receptor superfamily, member 5
• Gene Symbols: CD40
• Host Species: Mouse
• Purification Method: Affinity chromatography
• Quantity: 100 Tests
• Regulatory Status: RUO
• Primary or Secondary: Primary
• Gene ID (Entrez): 958
• Target Species: Human
• Content And Storage: 4°C, store in dark, DO NOT FREEZE!
• Product Type: Antibody
• Form: Liquid
• Isotype: IgG1 κ

Frequently Asked Questions (FAQs)

Can I use the OneComp and UltraComp eBeads microspheres with Super Bright-conjugated antibodies?

UltraComp eBeads microspheres (Cat. No. 01-2222) are recommended for use with Super Bright dyes.
Note: Super Bright Staining Buffer (Cat. No. SB-4400) is not compatible with UltraComp eBeads microspheres (Cat. No. 01-2222-41, 00-2222-42). If using UltraComp eBeads microspheres as a compensation tool, solely use Flow Cytometry Stain Buffer (Cat. No. 00-4222-26, 00-4222-57) for any antibody dilutions.

In some experiments, we have observed that compensation values for Super Bright 780- and Brilliant Violet 785- or Brilliant Violet 786-conjugated antibodies are higher in the violet 450/50 channel when using UltraComp eBeads microspheres as compared to single-color stained cells. In such circumstances, we would recommend setting compensation with cells. We have also observed this in some experiments using AbC Total Antibody Compensation beads, both with Super Bright 780 and Brilliant Violet 786. We have not tested Brilliant Violet 785 with the AbC beads.

Can I prepare an antibody cocktail containing Super Bright Staining Buffer and Super Bright-conjugated antibodies ahead of time?

We recommend that the antibody cocktails containing Super Bright-conjugated antibodies and Super Bright Staining Buffer are prepared fresh prior to staining. Discard any unused portions. We do not recommend overnight storage of prepared cocktails.

Can I fix my cells after staining with Super Bright-conjugated antibodies? How long can fixed cells be stored prior to analysis?

Samples that have been stained with antibodies conjugated to Super Bright dyes may be stored for up to three days, at 2-8°C, in the dark, using either IC Fixation Buffer (Cat. No. 00-8222) or 1-step Fix/Lyse Buffer (Cat. No. 00-5333) with no significant effect on brightness or compensation.

Are the Super Bright dyes sensitive to methanol fixation?

Super Bright dyes are stable in methanol-based fixation buffers.

Can Super Bright-conjugated antibodies be used in combination with intracellular (IC Fixation/Permeabilization) or intranuclear (Foxp3 fixation/permeabilization) staining?

Yes, Super Bright-conjugated antibodies are stable in formaldehyde-based fixation buffers and permeabilization buffers, such as the IC Fixation and Permeabilization Buffer Set (Cat. No. 88-8824) and the Foxp3/Transcription Factor Staining Buffer Set (Cat. No. 00-5523).

Which laser do I use to detect Super Bright-conjugated antibodies?

The violet laser (405 nm) should be used to excite Super Bright-conjugated antibodies.

Can the Super Bright Staining Buffer be used with other polymer dyes?

Yes, the Super Bright Staining Buffer (Cat. No. SB-4400) is compatible with other polymer dyes (i.e., Brilliant Violet dyes) and is useful for minimizing any non-specific polymer interactions when two or more of these dyes are used in combination.

Can Super Bright-conjugated antibodies be used in combination with other polymer dyes? What buffer should I use when using more than one Super Bright or polymer dye?

When using two or more Super Bright dyes, or when using Super Bright dyes in combination with other polymer dyes (i.e., Brilliant Violet dyes), we recommend using the Super Bright Staining Buffer (Cat. No. SB-4400) to minimize any non-specific polymer interactions between these fluorochromes. For specific instructions for use, please refer to the product technical data sheet.

Are there recommended tubes for staining with Super Bright-conjugated antibodies?

We recommend using polystyrene tubes (for example, FACS tubes) for staining with your Super Bright-conjugated antibodies. If using polypropylene (for example, Eppendorf tubes), protecting from light is critical.

Is there a specific buffer I should use when staining with Super Bright-conjugated antibodies?

No special buffer is required when using a single Super Bright-conjugated antibody in a panel.
When using more than one Super Bright dye, or when using Super Bright dyes in combination with other polymer dyes (i.e., Brilliant Violet dyes), we recommend using the Super Bright Staining Buffer (Cat. No. SB-4400) to minimize any non-specific polymer interactions between these fluorochromes. For specific instructions for use, please refer to the product technical data sheet.

How does the protocol for staining with Super Bright-conjugated antibodies differ from staining with conventional organic fluorochrome and eVolve conjugated antibodies?

Antibodies conjugated to Super Bright dyes can be used similarly to traditional fluorochromes. If multiple Super Bright dyes are used in combination with each other or in combination with other polymer dyes (i.e., Brilliant Violet dyes), then the use of Super Bright Staining Buffer (Cat. No. SB-4400) is recommended to minimize any non-specific polymer interactions between these fluorochromes. For specific instructions for use, please refer to the product technical data sheet.

Can the Super Bright dyes be used for non-flow applications?

Super Bright dyes have not been tested for applications other than flow cytometry.

What is the difference between the Super Bright and eVolve dyes?

Super Bright dyes are fluorochromes based on polymer technology and are excited by the violet laser. In contrast, eVolve dyes are based on Qdot nanocrystal technology and are maximally excited by the UV laser, although they may also be excited by violet, blue, yellow-green, and, depending on the eVolve, the red laser lines. The general shape of the emission spectra for Super Bright dyes will be more similar to traditional fluorochromes, while eVolve dyes have extremely narrow emission spectra. Please contact Tech Support (techsupport@thermofisher.com) for more information.

What are the Super Bright dyes?

eBioscience Super Bright dyes are a series of patent-pending fluorochromes that are based upon a fluorescent polymer and its tandems. Super Bright dyes have been developed for use in flow cytometry and can be excited by the violet laser (405 nm). The peak emission of each Super Bright dye is indicated by the number in the product description, e.g., "Super Bright 600" has a peak emission of 600 nm.

Are the Super Bright Dyes photo-labile?

As with other fluorochromes, we recommend minimal exposure to light to maintain optimal signal.

Safety and Handling

WARNING: Cancer - www.P65Warnings.ca.gov