CLIA was born in 1977. According to the basic principle of radioimmunoassay, chemiluminescence immunoassay was established by combining high sensitive chemiluminescence technology with high specific immune response. CLIA has the advantages of high sensitivity, strong specificity, wide linear range, simple operation and no need of expensive equipment.
CLIA is widely used in the detection of antigens, haptens and antibodies of different molecular sizes, as well as nucleic acid probes. Compared with RIA, IFA and EIA, CLIA has the advantages of non radiation, long validity period and full automation.
Chemiluminescence immunoassay consists of two systems: immunoassay and chemiluminescence analysis. Immunoassay system uses chemiluminescent substances or enzymes as markers, which are directly labeled on the antigen or antibody. After the reaction of antigen and antibody, antigen antibody immune complex is formed.
Chemiluminescence analysis system is after the end of the immune reaction, add oxidant or enzyme luminescent substrate, chemiluminescence material after oxidation by oxidant, form an excited state intermediate, will emit photons to release energy to return to the stable ground state, the luminous intensity can be detected by luminous signal measuring instrument. According to the relationship between chemiluminescent markers and luminous intensity, the content of the measured substance can be calculated by standard curve.
Chemiluminescent immunoassay (CLIA) is a kind of immunoassay method which uses chemiluminescent agent to label antibody or antigen directly. At present, luminol and acridine esters are the most common chemiluminescent agents.
1 chemiluminescence immunoassay of luminol: luminol is an oxidative reaction. Luminol can be oxidized by many oxidants in alkaline solution, among which H2O2 is the most commonly used. Due to the slow reaction rate, some enzymes or inorganic catalysts need to be added. The main enzymes are horseradish peroxidase (HRP), and the inorganic ones include O3, halogen, Fe3, Cu2, CO2 and their complexes.
In the early stage, it was mainly used for the determination of inorganic and organic biological small molecules, and the sensitivity decreased due to the decrease of the luminous intensity after labeling. It has been found that the addition of Some Phenols and their derivatives, amines and their derivatives, and phenylboronic acid derivatives can significantly enhance the luminescence of the system. The luminescence intensity can be increased by 1000 times, and the "background" luminescence is significantly reduced, and the luminescence time is also prolonged. The use of these enhancers makes chemiluminescence immunoassay widely used in protein and nucleic acid analysis.
2 acridine ester labeled chemiluminescence immunoassay: acridine ester was used in CLIA. Because of its poor thermal stability, more stable acridine ester derivatives were synthesized. Under the condition of H2O2 and oh -, acridine esters can emit light rapidly with high quantum yield. For example, the quantum yield of acridine aromatic esters can reach 0.05. When acridine esters are used as markers for immunoassay, the luminescence system is simple, rapid, and does not require the addition of catalyst. Moreover, the labeling efficiency is high and the background is low. These characteristics arouse the great interest of the majority of analysis and diagnosis workers.
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