High Sensitive Rat IgG ELISA Assay Kit

$550.00

The Eagle Biosciences High Sensitive Rat IgG ELISA Assay Kit is intended  for the quantification of Rat IgG in serum, plasma, cell culture supernatants. The Eagle Biosciences High Sensitive Rat IgG ELISA Assay Kit is for research use only and should not be used for diagnostic procedures.

High Sensitive Rat IgG ELISA Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 0.8 pg/ml
Dynamic Range: 1.56-100 pg/ml
Incubation Time: 3.5 hours
Sample Type: Serum, Plasma, Cell culture Supernatants
Sample Size: 100 µL

Additional Information

Assay Background

Antibodies are major components of the immune system. IgG is the main antibody isotype found in blood and extracellular fluid allowing it to control infection of body tissues. By binding many kinds of pathogens—representing viruses, bacteria, and fungi—IgG protects the body from infection. It does this via several immune mechanisms: IgG-mediated binding of pathogens causes their immobilization and binding together via agglutination; IgG coating of pathogen surfaces (known as opsonization) allows their recognition and ingestion by phagocytic immune cells; IgG activates the classical pathway of the complement system, a cascade of immune protein production that results in pathogen elimination; IgG also binds and neutralizes toxins. IgG also plays an important role in antibody-dependent cell-mediated cytotoxicity (ADCC) and intracellular antibody-mediated proteolysis, in which it binds to TRIM21 (the receptor with greatest affinity to IgG in humans) in order to direct marked virions to the proteasome in the cytosol. IgG is also associated with Type II and Type III Hypersensitivity. IgG antibodies are generated following class switching and maturation of the antibody response and thus participate predominantly in the secondary immune response. IgG is secreted as a monomer that is small in size allowing it to easily perfuse tissues. It is the only isotype that has receptors to facilitate passage through the human placenta, thereby providing protection to the fetus in utero. Along with IgA secreted in the breast milk, residual IgG absorbed through the placenta provides the neonate with humoral immunity before its own immune system develops. Colostrum contains a high percentage of IgG, especially bovine colostrum. In individuals with prior immunity to a pathogen, IgG appears about 24–48 hours after antigenic stimulation.

Assay Principle

This assay employs the quantitative sandwich enzyme immunoassay technique. An antibody specific for IgG has been pre-coated onto a microtiter plate. Standards or samples are pipetted into the wells and any IgG present is bound by the immobilized antibody. After washing away any unbound substances, a HRP-conjugated antibody specific for IgG is added to each well and incubate. Following a washing to remove unbound substances, a substrate solution (TMB) is added to the wells and color develops in proportion to the amount of IgG bound in the initial step. The color development is stopped by the addition of acid and the intensity of the color is measured at a wavelength of 450nm ±2nm.The concentration of IgG in the sample is then determined by comparing the O.D of samples to the standard curve.

Assay Procedure

1. Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.

2. Add 100 μl of standards, samples and zero controls (standard diluent buffer) into wells. Incubate for 2 h at 36 °C.

3. Aspirate each well and wash, repeating the process four times for a total five washes. Wash by filling each well with 1× Wash Buffer (350 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating, decanting or blotting against clean paper towels.

4. Add 100 μl 1X Antibody conjugate into each well. Cover wells and incubate for 60 min at 36°C.

5. Aspirate each well and wash as step 3.

6. Add 100 μl of TMB Reagent to each well. Incubate for 15 minutes at 36°C in dark.

7. Add 100 μl of Stop Solution to each well. The color of the solution should change from blue to yellow.

8. Read the OD with a microplate reader at 450nm immediately.

Manual

Product Manual