Invitrogen™ MHC Class II (I-A/I-E) Monoclonal Antibody (M5/114.15.2), Super Bright™ 645, eBioscience™, Invitrogen™

Catalog No.	Quantity
64532182	100 μg

Catalog No. 64532182 Supplier Invitrogen™ Supplier No. 64532182

Description

Rat Monoclonal Antibody

Description: This DS5MMER monoclonal antibody recognizes mouse MerTK, a 170-210 kDa member of the TAM family of tyrosine kinase receptors that also includes Axl and Tyro3. MerTK is expressed on tissue macrophages and is involved in the removal of apoptotic cells. This process relies on two soluble ligands of MerTK, Protein S and Gas6 that bind to phosphatidylserine found on the outer leaflet of the plasma membrane of cells undergoing apoptosis. Upon binding these ligands, MerTK undergoes autophosphorylation at multiple tyrosine residues that activate the PI3K and Akt pathways. This results in the phagocytosis of apoptotic cells and also results in the direct inhibition of TLR-induced production of pro-inflammatory cytokines. In addition, MerTK may function as a putative entry receptor for filoviruses. Deficiency of MerTK causes general autoimmunity, inflammation and accumulation of apoptotic bodies. MerTK is constitutively released from the cell surface by metalloproteinases and thus is present in the serum and culture medium. This process can be enhanced by stimulation with LPS. MerTK can be used to help discriminate macrophages from dendritic cells. MerTK is often expressed on maligt cells and may be implicated in immune evasion. Applications Reported: This DS5MMER antibody has been reported for use in flow cytometric analysis. Applications Tested: This DS5MMER antibody has been tested by flow cytometric analysis of resident peritoneal macrophages.

MHC (major histocompatibility complex) class II molecules are transmembrane glycoproteins expressed on the surface of professional antigen-presenting cells, such as macrophages, dendritic cells and B cells. Before their exposition on the cell surface, the MHC class II molecules react with endocytosed exogenous antigens, which are then presented to the T cells. The antigen-binding grove between MHC class II alpha and beta chain is open at both ends and is 15-24 amino acid residues long. HLA and MHC antibodies play a significant role in Immunopeptidomics, facilitating the identification and characterization of neoantigens through high-performance liquid chromatography coupled to tandem Mass Spectrometry.

Specifications

• Antigen: MHC Class II (I-A/I-E)
• Applications: Flow Cytometry
• Classification: Monoclonal
• Clone: M5/114.15.2
• Concentration: 0.2 mg/mL
• Conjugate: Super Bright 645
• Formulation: PBS with BSA and 0.09% sodium azide; pH 7.2
• Gene: H2-Ab1
• Gene Accession No.: P01910, P01921, P04227, P04228, P04230, P06342, P06343, P06344, P06345, P06346, P14434, P14435, P14436, P14437, P14438, P14483, P23150
• Gene Alias: Abeta; AI323765; AI845868; cell surface glycoprotein; E-alpha-f; H-2 class II histocompatibility antigen, A beta chain; H-2 class II histocompatibility antigen, E-D alpha chain; H-2 class II histocompatibility antigen, E-K alpha chain; H-2 class II histocompatibility antigen, E-U alpha chain; H-2 class II histocompatibility antigen, I-E beta chain; H-2Ab; H2-Ab; H2-Ab1; H-2Ea; H2-Ea; H2-Ea-ps; H2Eb; H-2Eb; H2-Eb1; H2-iabeta; H2-IE-alpha; histocompatibility 2, class II antigen A, beta 1; histocompatibility 2, class II antigen E alpha; histocompatibility 2, class II antigen E alpha, pseudogene; histocompatibility 2, class II antigen E beta; Ia2; Ia-2; Ia3; Ia-3; Ia4; Ia-4; IAb; I-Ab; I-Abeta; I-Ad; I-Aq; I-E alpha MHC class II; I-E beta MHC class II; I-Ealpha; I-Ed; I-Ek; major histocompatibility complex class II beta chain; major histocompatibility complex H-2E beta chain; MHC class 2; MHC class II antigen A beta; MHC class II antigen E alpha; MHC class II H2-IA-beta-psi; MHC class II IE-b; MHC class II protein; MHC H2-IE-beta cell surface glycoprotein; response to metastatic cancers 1; Rmcs1
• Gene Symbols: H2-Aa, H2-Ab1, H2-Eb1
• Host Species: Rat
• Purification Method: Affinity chromatography
• Quantity: 100 μg
• Regulatory Status: RUO
• Primary or Secondary: Primary
• Gene ID (Entrez): 14960, 14961, 14969
• Target Species: Mouse
• Content And Storage: 4°C, store in dark, DO NOT FREEZE!
• Product Type: Antibody
• Form: Liquid
• Isotype: IgG2b 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.

For Research Use Only