Polyclonal antibodies injection

When an antigen is injected into an animal, the resulting antibodies are polyclonal, being synthesized by a mixture of B cells. Polyclonal antibodies are directed against a number of different sites (epitopes or determinants) on the antigen and thus are not monospecific. However, by means of a method developed by Kohler and Milstein, large amounts of a single monoclonal antibody specific for one epitope can be obtained. 

Polyclonal Antibodies against FORL r purified polygalacturonase were raised in white rabbits. For the first immunization 200 jxg of purified protein in 300 jxl of distilled water was mixed with 200 1 of PBS and 500 n of complete Freund s adjuvcmt and injected intramusculary into the leg. Two subsequent intramuscular injections, each containing 300 fig of protein in 1 ml of incomplete Freund s adjuvant were given at 1 month intervals. Finally, the rabbit was bled 1 week later. The antisera, separated from blood by incubation at 37 "C, were stored in 1 ml fractions at -20 C. 

Fig. 20.2 Chronic lithium and valproate robustly increase bcl-2 immunoreactive neurons in the frontal cortex. Male Wistar Kyoto rats were treated with either Li2C03, valproate or saline by twice daily i.p. injections for four weeks. Rats brains were cut at 30 pm serial sections were cut coronally through the anterior portion of the brain, mounted on gelatin-coated glass slides and were stained with thio-nin. The sections of the second and third sets were incubated free-floating for 3 d at 4°C in 0.01 M PBS containing a polyclonal antibody against bcl-2 (N-19, Santa Cruz Biotechnology,...

Generation of antibodies that can recognize and bind to specific viruses is straightforward. A sample of live or attenuated virus, or a purified component of the viral caspid, can be injected into animals to stimulate polyclonal antibody production (or to facilitate monoclonal antibody production by hybridoma technology). Harvested antibodies are then employed to develop specific immunoassays that can be used to screen test samples routinely for the presence of that specific virus. Immunoassays capable of detecting a wide range of viruses are available commercially. The sensitivity, ease, speed and relative inexpensiveness of these assays render them particularly attractive. 

Polyclonal antibody preparations have been used for several decades to induce passive immunization against infectious diseases and other harmful agents, particularly toxins. The antibody preparations are usually administered by direct i.v. injection. While this affords immediate immunological protection, its effect is transitory, usually persisting for only 2-3 weeks (i.e. until the antibodies are excreted). Passive immunization can be used prophylactically (i.e. to prevent a future medical episode) or therapeutically (i.e. to treat a medical condition that is already established). An example of the former would be prior administration of a specific anti-snake toxin antibody preparation to an individual before they travel to a world region in which these snakes are commonly found. An example of the latter would be administration of the anti-venom antibody immediately after the individual has experienced a snake bite. 

Polyclonal antibodies are produced by injecting an antigen into an animal in the presence of an adjuvant containing bacterial lipopolysaccharides that stimulate the immune system. Serum prepared from the blood contains several different classes of antibodies that interact with different domains in the antigen molecule, each of... 

To produce monoclonal antibodies, the first antigen injection in mice is followed a few weeks later by a booster of the same antigen. When polyclonal antibodies are detected in the serum of mice a few days after the booster injection,... 

In 1895, Hericourt and Richet reported the first clinical trials testing the principle of antibody production. They injected cancer cells into animals to obtain antiserum to treat cancer patients this was the first time several patients with cancer were administered tailor-made serum for treatment. Several patients showed improvement, which was encouraging, but none of the patients were completely cured. These trials were repeated in the early 1900s but the results were not consistent. The problems included the variability of the antisera and the side effects of polyclonal antibodies - some of which were directed against self. 

The immunisation procedure (at least 12 weeks with booster injections, addition of oil based adjuvants to stimulate the immunogenic response) gives rise to an antiserum representing a mixture of polyclonal antibodies which are produced by different immunocompetent cells and thus have different binding sites and affinities. 

Example 1. A rabbit has been injected with a protein with the intent of generating polyclonal antibodies to the immunogen. After adequate time for the generation of the immune response, a blood sample is drawn from the rabbit and serum is analyzed for the presence of the antibodies by ELISA (Figure 7.6). 

Figure 6 illustrates the progressive overloading of an anti-HSA polyclonal antibody column after repeated injections of 2 /zg of HSA. At first injections, impurities elute from the column at the dead volume, while HSA is totally adsorbed. The gradual emergence of the nonretained HSA elution peak is due to two different effects, the saturation of the support and the slow adsorption kinetic process. The unretained fraction is calculated from peak area measurements, subtracting the area of the impurity response peak. 

Polyclonal Antibodies After an antigen is injected into an animal by a regimen designed to induce an optimal immune response, serum can be collected from the animal and the immunoglobulin fraction isolated. This antisera is enriched with antibodies specific for the original antigen. Because a large number of lymphocytes are involved in the production of the antisera, antibodies produced by this classical method are called polyclonal. 

The polyclonal antibodies obtained by immunizing experimental animals have been and will continue to be satisfactory reagents for many immunoassay methods. Choices can be made among adjuvants, routes of injection, dosage and immunization schedules, species of animal to be immunized, etc. (2-6, 28-31). 

---