Biotechnology Drug Analysis Method：Ligand Binding Test

After ingesting biotechnology drugs, the body will produce different degrees of immune response based on the immunogenicity of the drug. The drug stimulates the body to form specific anti-drug antibodies (ADA) for the immunogenicity of the drug, allergies, autoimmunity and different The pharmacokinetic characteristics of the drug are anti-drug antibody reactions, which can cause various clinical adverse reactions such as drug neutralization, abnormal biodistribution, and enhanced drug clearance, which may change the efficacy of the drug. Ligand binding test is the main and commonly used bioanalytical method for biotechnology drugs. Today we will briefly understand this method.

Biotechnology drugs can cause a certain anti-drug antibody response, that is, immune response. The body’s immune response to biological drugs may affect the safety and effectiveness of the product, but the clinical effect of the immune response is highly variable, and some are completely healthy No impact, some will cause extremely harmful effects. Neutralizing antibody refers to a type of anti-drug antibody that can directly bind to the drug binding site or through steric hindrance, so that the drug loses its ability to bind to its target, thereby blocking and neutralizing drug treatment Active antibodies. Neutralizing antibodies can affect the safety and effectiveness of the drug by affecting the exposure level of the drug in the body. The ligand binding test can be used to evaluate the neutralizing antibody. The ligand binding test methods mainly include enzyme-linked immunosorbent assay, radioimmunoassay, and time-resolved fluorescence immunoassay.

Ligand binding test method Incubate the radiolabeled antibody and over-shooting unlabeled ligand with the receptor to be tested at the same time, determine the amount of labeled ligand in the bound and free state, and calculate the maximum number of binding sites and dissociation constant for the receptor to be tested Methods of analysis. Medicilon Bioanalysis Department can provide FDA/CFDA GLP-compliant macromolecular drug bioanalysis services to support the screening and development of protein drugs, antibody drugs, vaccines and biomarkers, as well as preclinical and clinical research.

(1) Enzyme-linked immunosorbent assay

Enzyme-linked immunosorbent assay, also called ELISA, can detect the content of antigen or antibody in the specimen. The basic method is to adsorb the known antigen or antibody on the surface of a solid-phase carrier (polystyrene microreaction plate) to make the enzyme-labeled antigen The antibody reaction is carried out on the surface of the solid phase, and the free components in the liquid phase are washed away by the washing method. In the development of biotechnology drugs, this technology can be used to detect macromolecular antigens and specific antibodies, etc., and has the advantages of rapid, sensitive, simple, and easy to standardize carriers. According to the connection and application form of the immunosorbent, conjugate and the test substance during the experiment, ELISA mainly includes sandwich method, indirect method, competition method and other types.

(2) Radioimmunoassay

The immune response caused by drugs is an important indicator of drug safety and effectiveness, which is also the common concern of regulatory agencies, manufacturers, clinicians and patients. Radioimmunoassay is also a method for analyzing the immunogenicity of antibody drugs. This method uses the sensitivity and accuracy of radionuclide detection and the specificity of antigen-antibody reaction combined with an immunological technique, including the principle of competitive binding reaction Radioimmunoassay (RIA) and non-competitive combined immunoradioassay (IRMA) are two methods. Radioimmunoassay technology is often used for the determination of trace substances such as various hormones, trace proteins, tumor markers and drugs.

(3) Time-resolved fluorescence immunoassay

Time-resolved fluorescence immunoassay (TRFIA) is a non-isotopic immunoassay technique developed on the basis of fluorescence analysis (FIA). This method uses lanthanide elements to label antigens or antibodies. According to the luminescence characteristics of lanthanide chelates, time-resolved technology is used to measure fluorescence, and the two parameters of wavelength and time are simultaneously detected for signal resolution, which can effectively eliminate non-specific fluorescence interference . It can be used to analyze hormones, viral hepatitis markers, tumor-associated antigens, pepsinogen (PG), drugs and peptides.

The above-described methods are relatively common methods for analyzing the immunogenicity of biotechnology drugs. In drug development, preclinical and clinical research, the establishment of effective and feasible anti-drug antibody detection methods can provide a powerful means for evaluating the immunogenicity of drugs.

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