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An Immunoassay Technique Based on Antigen-Antibody Reaction-RIA Immunoassay

2020-12-03
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Biopharmaceutical analysis can detect and study the quality of various biological drugs. Radioimmunoassay (RIA) is a method of biopharmaceutical analysis. This method is based on the basic principle that the radionuclide-labeled antigen competes with the unlabeled antigen in the reaction system to bind to specific antibodies to determine the amount of antigen in the sample to be tested. It is often used for various hormones, trace proteins, tumor markers, drugs and other trace substances The determination is especially suitable for the ultra-micro analysis of hormones, peptides and other small substances.   ELISA is a method of biopharmaceutical analysis. It is a more general analysis method in the field of immunoassay. This method uses enzyme-labeled antigen or enzyme-labeled antibody as the main reagent, and the color of the enzyme-catalyzed substrate in the complex is The qualitative or quantitative labeling and immunoassay technology for substances has the advantages of simple operation, fast, sensitive, economical and suitable for batch processing.   Different from ELISA, radioimmunoassay technology uses radioisotopes to label antigens or antibodies, and analyzes samples through separation of reactants and detection of radioactive signals. However, RIA immunoassay technology has the problem of attenuation of isotope signals and can cause harm to the human body during the detection process, so this technology is now rarely used. Let’s take a look at the main steps of RIA.

(1) Antigen-antibody reaction     

According to the principle of RIA, unlabeled antigens (standards and samples to be tested), labeled antigens and specific antibodies are added to the reaction tube, and the competitive inhibition reaction is carried out under certain conditions (temperature, time and pH of the medium).    

(2) Separate bound and free markers     

After the RIA reaction balances, the labeled antigen and the reagent antibody form an immune complex (B). Because its content is very small, it cannot precipitate by itself, so it needs to be added with a suitable precipitant to completely precipitate it, and then centrifuged to separate it from the free labeled antigen (F). Some small molecule antigens can also be separated from B and F by adsorption.

(3) Radioactivity measurement and data processing     

After separating B and F, the radioactivity of the labeled antigen-antibody complex (B) can be measured, and the free labeled antigen (F) can also be determined according to the RIA method and purpose. Draw a standard curve (dose-response curve). The sample tube uses its measured or calculated reaction parameters to find out the corresponding antigen concentration to be tested through the standard curve. At present, computers are commonly used for data processing, automatic drawing and printing of standard curves Sample antigen concentration.

Although the radioimmunoassay method is easy to operate and low in cost, it reached its peak in the late 1970s, but due to its radioactivity, difficult to store and difficult to automatically detect and other defects, its use was limited, and began to decline and replace in the late 1970s. However, no immunoassay technique is perfect at present, and various techniques need to be continuously developed and improved to develop newer and more ideal immunoassay techniques.

Medicilon Bioanalysis department has a full range of multifunctional technology platforms such as SpectraMaxM4/M5/i3x, MSD, Luminex, Biacore 8K, Envision, Gyrolab, ABI7500 qPCR, etc. It uses the LIMS system to implement comprehensive laboratory information management, providing Biotechnology drug analysis services that fully comply with FDA/CFDA/OECD GLP specifications.

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