Rat Pure-Blot anti-Mouse IgG (H+L) Secondary Antibody (eB144) [Biotin]

• Reactivity: Mu
• Applications: WB, ELISA
• Clone: eB144
• Clonality: Monoclonal
• Host: Rat
• Conjugate: Biotin

Rat Pure-Blot anti-Mouse IgG (H+L) Secondary Antibody (eB144) [Biotin] Summary

• Description: This secondary antibody was prepared from tissue culture supernatant by Protein G affinity chromatography. Assay by Immunoelectrophoresis resulted in a single precipitin arc against Anti-Mouse Serum.
  
  Store vial at 4C before opening. DO NOT FREEZE. This product is stable at 4C as an undiluted liquid. Dilute only prior to immediate use.
• Immunogen: Mouse IgG
• Isotype: IgG
• Clonality: Monoclonal
• Host: Rat
• Purity: Protein G purified

Applications/Dilutions

• Dilutions:
  • ELISA
  • Western Blot 1:000
• Application Notes: This antibody is ideal for use in protocols involving immunoblotting of immunoprecipitated proteins. This antibody preferentially detects the non-reduced form of mouse IgG (IgG1, IgG2a, IgG2b, IgG3) over the reduced, SDS-denatured form of IgG. To achieve best results when detecting mouse IgG1 subtypes, we recommend performing a dot blot or titration to determine the optimal dilution factor for your desired application. When the immunoprecipitate is fully reduced immediately prior to SDS-gel electrophoresis, reactivity of This antibody with the 55 kDa heavy chains and the 23 kDa light chains of the immunoprecipitating antibody is minimized thereby eliminating interference by the heavy and light chains of the immunoprecipitating antibody in IP/Western blot applications. Applications include studies examining post-translational modification (e.g., phosphorylation or acetylation) or protein-protein interactions. Note: There are two key procedural considerations: 1. When using any Pure-Blot reagent, ensure there is complete reduction of the lysate. 2. Use BLOTTO/milk powder for complete and effective blocking of the western blot. All recommended dilutions for listed applications are intended as an initial recommendation, specific conditions for each protein and antibody combination should be specifically optimized by the end user.

Packaging, Storage & Formulations

• Storage: Store at 4C. Do not freeze.
• Buffer: 0.01 M Sodium Phosphate, 0.15 M Sodium Chloride, pH 7.2 (Proprietary Stabilizer)
• Preservative: 0.09% Sodium Azide
• Purity: Protein G purified

Alternate Names for Rat Pure-Blot anti-Mouse IgG (H+L) Secondary Antibody (eB144) [Biotin]

• IP Detection Reagent

Background

Antibodies, also known as immunoglobulins (Igs) are critical for immunity and are grouped into five primary classes: IgG, IgM, IgA, IgD, and IgE. The most abundant antibody isotype is immunoglobulin G (IgG) with concentrations ranging from 7.5-22 mg/ml in human serum and has a molecular weight of 150 kDa. The major effector functions of IgG include neutralization, opsonization, complement fixation and antibody dependent cell-mediated cytotoxicity (ADCC). This monomeric immunoglobulin, expressed on the surface of mature B cells, is often depicted as a Y-shape and comprised of 2 heavy chains and 2 light chains linked by disulfide bonds. The heavy chain is type gamma including subtypes gamma 1, gamma 2, gamma 3, and gamma 4 while the light chain is either a kappa or lambda chain. An IgG molecule has two antigen binding sites, each consisting of a heavy and light chain N-terminal variable domain. When combined with the constant heavy chain 1 (Ch1) and the constant light chain domains, it forms the fragment antigen-binding (Fab) region (2 per antibody). The remaining domains (Ch2-Ch4) of both heavy chains make up the Fc region and contain a site for covalently linking an enzymatic or fluorochrome probe, such as HRP or Janelia Fluor 549, for target detection and visualization (1,2,3).

The 4 IgG subclasses, sharing 95% amino acid identity, include IgG1, IgG2, IgG3, and IgG4 for humans and IgG1, IgG2a, IgG2b, and IgG3 for mice. The relative abundance of each human subclass is 60% for IgG1, 32% for IgG2, 4% for IgG3, and 4% for IgG4. In an IgG deficiency, there may be a shortage of one or more subclasses (4).

References

1. Painter RH. (1998) Encyclopedia of Immunology (Second Edition). Elsevier. 1208-1211

2. Chapter 9 - Antibodies. (2012) Immunology for Pharmacy. Mosby 70-78

3. Schroeder H, Cavacini, L. (2010) Structure and Function of Immunoglobulins. J Allergy Clin Immunol. 125(2 0 2): S41-S52. PMID: 20176268

4. Vidarsson G, Dekkers G, Rispens T. (2014) IgG subclasses and allotypes: from structure to effector functions. Front Immunol. 5:520. PMID: 25368619

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Secondary Antibodies are guaranteed for 1 year from date of receipt.