Since the ELISA method is based on the immunological reaction between the antigen and the antibody, it is specific. On the other hand, since the enzyme-labeled antigen or antibody is a combination of an enzyme molecule and an antigen or an antibody molecule, it can catalyze the reaction of the substrate molecule and produce amplification, which is because of this amplification. Sensitivity. Therefore, ELISA is a sensitive and specific method.
ELISA can be used to determine antigens and can also be used to determine antibodies. There are three essential reagents in this assay:
(1) a solid phase antibiotic pro- or antibody, ie an "immunosorbent";
(2) an enzyme-labeled antigen or antibody, referred to as a "conjugate";
(3) Substrate for the enzyme reaction.
Depending on the source of the reagent and the condition of the specimen and the specific conditions of the assay, various types of assays can be devised. There are several types of ELISAs for clinical testing:
Double antibody sandwich assay
The double antibody sandwich method belongs to the non-competitive binding assay. It is the most commonly used ELISA for detecting antigens and is suitable for detecting multivalent antigens with at least two antigenic determinants in the molecule, but not for detection of small molecule haptens.
The basic working principle is that the antibody and the enzyme-labeled antibody linked to the solid phase carrier are respectively combined with two antigenic determinants on the detected antigen molecule in the sample to form a solid phase antibody-antigen-antibody standard antibody immune complex. Since the amount of the solid phase antibody and the enzyme-labeled antibody in the reaction system is excessive relative to the antigen to be tested, the amount of the complex formed is proportional to the content of the antigen to be tested (within the detectable range of the method). The mass of the colored substance (OD value) generated by the enzyme in the complex acting on the added substrate can be determined to determine the antigen content to be tested.
If the antibody and the enzyme-labeled antibody on the solid phase carrier are respectively combined with two different antigenic determinants on the detected antigen molecule in the sample, it belongs to the two-site sandwich method.
If the antigen on the solid phase carrier and the enzyme-labeled antigen are respectively bound to the detected antibody molecule in the sample, it is a double antigen sandwich method.
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