Monoclonal antibody technology is an immunological technique in which a single B lymphocyte producing an antibody is hybridized with a bone marrow tumor cell to obtain a hybrid cell which can produce both an antibody and an unlimited proliferation, and thereby produce an antibody. An antibody produced by only one type of cell, corresponding to a polyclonal antibody, a plurality of antibodies, an antibody produced by various types of cells.
Processes
(1) Preparation of antigen.
The immunological antigen for preparing a monoclonal antibody is not required to be high in purity, but the high-purity antigen increases the chance of obtaining the desired monoclonal antibody and at the same time reduces the workload of screening. Therefore, the purer the immune antigen, the better, and should be determined based on the antigen being studied and laboratory conditions. In general, the source of the antigen is limited, or the nature is unstable, it is apt to be denatured during purification, or its immunogenicity is strong, or the desired monoclonal antibody is used for purification or analysis of different components of the antigen, and the antigen for immunization is only Initial purification is not required, but there are many impurities in the antigen, especially if the immunogenicity of these hybrids is strong, the antigen must be purified. The antigen for detection may be of the same purity as the immunizing antigen, or may be of different purity depending on the type of screening method and its specificity and sensitivity.
(2) Selection of immunized animals.
Mice and rats can be used as immunized animals depending on the myeloma cells used. Because all mouse myeloma cell lines for hybridoma technology are derived from BALB/c mice, all rat myeloma cells are derived from LOU/c rats, so the general hybridoma production is used. These two pure animals are used as immunized animals. However, sometimes interspecific hybridization is required for special purposes, and other animals can be immunized. Interspecies hybridomas generally have an unstable ability to secrete antibodies because chromosomes are easily lost. In the case of mice, it is preferred to use 8-12 weeks of age for the first immunization, and female rats are more convenient to handle.
(3) Determination of the immunization procedure.
Immunization is one of the important steps in the preparation of monoclonal antibodies. The purpose is to differentiate and proliferate B lymphocytes under the stimulation of specific antigens, so as to facilitate cell fusion to form hybrid cells and increase the chance of obtaining hybridomas secreting specific antibodies. Therefore, in designing the immunization program, the nature and purity of the antigen, the amount of antigen, the route of immunization, the number and timing of immunizations, the application of the adjuvant, and the ability of the animal to respond to the antigen should be considered. No immunological procedure can be applied to a variety of antigens. Most of the current immunization procedures are based on methods for preparing conventional polyclonal antibodies. Table 6-1 lists the currently used immunization procedures. Immunization pathways commonly used in vivo immunoassays include subcutaneous injections, intraperitoneal or intravenous injections, as well as footpads, intradermal, nasal drops or eye drops. The last booster is usually administered intraperitoneally or intravenously, and the latter is especially recommended because it allows the antigen to act more rapidly and fully on spleen cells. Spleen fusion was good on the third day after the last booster immunization. The results of many laboratories showed that in the primary immunization and re-immune response, the spleen cells were fused with myeloma cells, and the peaks of specific hybridomas were respectively On days 4 and 22, the hybridomas obtained at the time of the primary immune response mainly secreted IgM antibodies, and the hybridomas obtained in the re-immunization response mainly secreted IgG antibodies. The authors realized that there was no significant parallel relationship between the peak of positive hybridoma and the titer of serum antibody in mice, and it was mostly before the peak of serum antibody. Therefore, in order to achieve the highest rate of hybridoma formation, it is necessary to have as many plasmablasts as possible, which is preferable to spleen for fusion on the third day after the last booster immunization. It has been reported that intrasplenic immunization can increase the immunoreactivity of mice to antigens and save time. Generally, it can be fused after 3 days of immunization.
Characteristics
Specificity
It is highly specific for a specific single antigenic epitope. Studies of anti-tumor antibody drugs have shown that its specificity is mainly characterized by specific binding, selective killing of target cells, targeted distribution in vivo, and Has a stronger effect.
Diversity
It is mainly reflected in the diversity of target antigens, the diversity of antibody structures, and the diversity of mechanism of action.
The third is directionality, and antibody drugs can be produced in a targeted manner, that is, antibody drugs having different therapeutic effects can be prepared as needed. Based on these characteristics, we can use it to make "biological missiles" to transport drugs to diseased parts, mainly cancer cells, to achieve therapeutic effects.
Clinical application
Disease diagnosis
The diagnosis of diseases using monoclonal antibodies is currently mainly manifested in the diagnosis of human diseases and livestock diseases, especially in the diagnosis of some infectious diseases and tumors. It is mainly diagnosed whether a person is infected with a corresponding disease by identifying a pathogen or a tumor antigen.
Disease treatment
At present, great progress has been made in the treatment of diseases with monoclonal antibodies, mainly by coupling the monoclonal antibodies with the drugs and then combining them with the specific antigens of the pathogens or tumors.
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