Overview of IgG Antibodies and Anti-human IgG Antibodies

What is an IgG antibody?

The IgG antibody is one of the types of mammalian antibodies (also known as immunoglobulins, Ig for short). The antibody-associated immunity against pathogen invasion is mainly provided by four types of antibodies, and is the only one that can provide passive immunity to the fetus through the placenta.IgG antibodies act to activate complement and neutralize multiple toxins in the immune response. IgG antibodies last a long time and are the only antibodies that protect the fetus from the placenta during pregnancy. They also secrete from the mammary gland into the colostrum, giving the newborn the first time to get antibody protection. IgG is a four-chain monomer, accounting for 75% of the total serum Ig. It is the most important antibody component in serum and extracellular fluid. Human IgG can be divided into four sub-categories, depending on the concentration in serum. IgG1, IgG2, IgG3, IgG4. IgG is synthesized three months after birth and is close to the adult level from three to five years old. It is mainly produced by plasma cells in the spleen and lymph nodes. It has a long serum half-life of about 20~23 days. It is the main antibody produced by the re-humoral immune response. It has high affinity, is widely distributed in the body, has important immune effects, and is anti-antibody. The main force of infection, so the clinical use of gamma globulin for immunotherapy should be injected every 2 to 3 weeks is appropriate.

IgG1, IgG3, and IgG4 can cross the placental barrier and play an important role in anti-infective immunity in neonates; IgG1, IgG2, and IgG4 can bind to staphylococcal protein A (SPA) through its Fc segment, thereby purifying antibodies, or For immunodiagnosis; IgG1, IgG3 can efficiently activate complement, and can bind to macrophage and NK cell surface Fc receptors, play a role in opsonization, ADCC, etc.; some autoantibodies and antibodies that cause type II and type III hypersensitivity reactions also belongs to IgG.

IgG antibody function:

It plays an important role in anti-infective immunity, especially in re-immune responses. IgG-type autoantibodies participate in type II and type III hypersensitivity reactions.

  1. Activate the classical pathway of complement, mediating bacteriolysis and cytotoxicity;
  2. Mediating the ADCC effect;
  3. Conditioning phagocytosis;
  4. Combine SPA;
  5. Neutralize toxins and viruses.

What are the techniques for IgG antibodies?

  1. Extraction of globulin

Mostly, ammonium sulfate salting out or sodium sulfate salting out is used. Ammonium sulfate salting has to be precipitated several times, with 40% saturation for the first time, 35% saturation for the second time, and 33% saturation for the third time. The gamma globulin after three extractions is basically an IgG component. The sodium sulphate method is simpler and the gamma globulin can be precipitated with 20%. Although the gamma globulin after salting out is mostly IgG, there are 5% other zone proteins, such as the gamma region heteroprotein. Interference is also caused by the inclusion of other so-called normal IgGs in the IgG component. Therefore, the γ-globulin crudely extracted by the salting-out method can only be used for general experiments or anti-serum with higher antibody titer.

  1. Ion exchange chromatography for IgG extraction

Commonly used ion exchangers are DEAE cellulose or QAE cellulose, which is ideal for QAE-Sephadex, and DE22, 32, 52 are also applicable. Take QAE-Sephadex A25 or A50 acid-treated and equilibrate in 0.05mol/L phosphate buffer pH 7.5-8.6, drain the water, weigh the wet weight Ig in 10ml serum, centrifuge at room temperature for 30min, filter or filter The ion exchanger is removed. The supernatant is treated once more to obtain a relatively pure IgG, even without other miscellaneous proteins. Purification of IgG by this technique is simple and does not damage the antibody, and can be extracted in small amounts or in large quantities.

  1. Affinity chromatography to extract specific IgG

The purified antigen or the crude antigen is cross-linked to Sepharose 4B to prepare an affinity chromatography column. After the antiserum is passed through the column, the unbound heteroprotein is washed away, and then eluted with potassium thiocyanate, and the pure specific IgG antibody is eluted. Because potassium thiocyanate has a destructive effect on the antibody, it should be dialyzed and removed in time. Purified IgG has low content and loses its protective effect. It should be applied in time or lyophilized. It can also be stored at -20 °C, but it is not ideal. Adding triethanolamine or glycerol can protect it.

  1. Preparation of F2 fragment by enzymatic hydrolysis

The point of action of pepsin on IgG is at the C-terminus (232 amino acids) of the disulfide bond connecting the two heavy chains, and as a result, the two Fabs are linked by a disulfide bond, retaining the binding site of the antibody. Compared with IgG, F(ab’)2 is characterized by the removal of the Fc segment, which eliminates the receptor action in cellular immunization experiments; it also causes IgG to lose its primary antigenic properties and is not bound by anti-IgG antibodies; indirect hemagglutination, sensitization of sheep red blood cells with F(ab’)2 is better than IgG.

Anti-human IgG antibody research

In the literature “Experimental study of anti-human IgG antibody combined with mitomycin in the treatment of bladder cancer”, in vitro experimental methods and tumor-bearing nude mice experimental methods were used to observe the combined use of anti-human IgG antibody and mitomycin C (MMC). The biological effects of inhibiting the growth of bladder cancer T24 cells and inducing apoptosis of tumor cells. As a result, it was found that, in addition to B lymphocytes and plasma cells, a variety of epithelial-derived tumor cells and some normal epithelial cells can produce IgG and have a function of promoting tumor cell growth. Anti-human IgG antibody can inhibit the growth of tumor cells, and can also significantly promote the apoptosis of tumor cells. In previous studies, we found that bladder cancer cells can express IgG significantly. Immunohistochemistry, mRNA in situ hybridization, RT-PCR and Western blotting methods also confirmed the presence of IgG in bladder cancer cell lines BIU-87 and T24.