Analysis Note:
The total protein is measured using the Biuret procedure with bovine albumin as the standard. To ensure that the product titer falls within the required range, antibody titer is standardized by microtiter plate ELISA with mouse IgG and human IgG. The product is tested for purity and specificity at final concentration by immunoelectrophoresis. The antibody is predominantly sheep IgG; no trace of albumin is detected.
Preparation Method:
Antibody and highly purified HRP (Rz >3.0) are conjugated under defined conditions to obtain optimally labeled product. Conjugated protein is purified by salt fractionation. The product is dialyzed into 0.02M sodium phosphate, 0.14M sodium chloride, pH 7.3, with 0.01% thimerosal, adjusted to standard titer, filtered through a 0.22 μm filter, vialed and lyophilized.
Background Information:
Immunoglobulin G (IgG) is a glycoprotein antibody that regulates immune responses such as phagocytosis and is also involved in the development of autoimmune diseases . Mouse IgGs have four distinct isotypes, namely, IgG1, IgG2a, IgG2b, and IgG3. IgG1 regulates complement fixation in mice. Anti-Mouse IgG (whole molecule)-Peroxidase antibody is specific for mouse IgG. Mouse IgG is a plasma B cell derived antibody isotype defined by its heavy chain. IgG is the most abundant antibody isotype found in mouse serum. IgG crosses the placental barrier, is a complement activator and binds to the Fc-receptors on phagocytic cells. The level of IgG may vary with the status of disease or infection. Horseradish Peroxidase (HRP) is an enzyme that catalyzes the conversion of chromogenic substrates such as o-phenylenediamine (OPD), 4-chloro-1-naphthol 3,3’,5,5’-tetramethylbenzidine (TMB), 3,3′-Diaminobenzidine (DAB) or 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS); chemiluminescent substrates such as CPS-3 (enhanced luminal) and fluorogenic substrates into detectable chromophores, light-emitters or fluorescers, respectively.
Applications:
Peroxidase conjugated Sheep IgG fraction to mouse IgG is used as a reagent in enzyme immunoassays (EIA), cell and tissue staining (for light microscopy), cell and tissue labeling (for electron microscopy), and blot immunostaining. (Note: F(ab')2 fragments are recommended for staining of cells or tissues which contain Fc receptors. Affinity purified antibodies or their fragments are recommended to avoid non-specific binding from inherent antibodies of host animals). It may be used to detect and quantitate the level of IgG in mouse serum and biological fluids via chromogenic, chemoluminescent or fluorogenic immunochemical or immunohistochemical techniques. It may also be used as a secondary antibody in assays that use mouse IgG as the primary antibody. The product can also be used for direct ELISA.
Product Description:
Product is the lyophilized powder of horseradish peroxidase (HRP)-conjugated sheep IgG fraction to mouse IgG (no cross to human) and buffer salts.
Recommended Use:
Reconstitution: Reconstitute product with 2.0 mL of deionized or distilled water. Gentle swirling may be used to speed rehydration. Do not add sodium azide. Avoid vigorous shaking of the reconstituted material.