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Biotin and avidin are currently used. In recent years, streptavidin (SA) has been used in the detection of BAS. It is a secretion in the culture of Streptomyces, intact streptavidin and extracted from egg white. Like avidin, it also consists of 4 identical peptide chains. Because streptavidin has a lower non-specific binding in detection applications than avidin, it has received increasing attention and has a tendency to replace avidin. Various antibodies or enzyme markers have been supplied in domestic products and are convenient to use.
BAS ELISA has the following characteristics:
Now they are introduced as follows:
Have you learned about bioavidin ELISA in your system?
Biotinvidin system (BAS) The technique of labeling antibodies is a new immunoassay developed in the late 1970s. Due to the strong affinity between avidin and biotin, the binding is rapid and extremely stable, the biotin-labeled antibody and enzyme have high labeling rate without affecting the activity of the protein, making the BAS labeling technique more conventional than enzyme-linked immunoassay and radioimmunoassay. And fluorescent immunoassay has higher sensitivity, which opens up a new way for the detection of trace antigens and antibodies.
1 for all antigen-antibody systems, including immunocytochemistry, ELISA, immunoblotting, etc., for a wide range of applications;
2BAS can bind to various biomacromolecules such as proteins, lipopolysaccharides, etc. under mild conditions, and this combination has no effect on the biological activity of protoplast macromolecules;
3 Each avidin molecule can bind to four biotin molecules, so that more enzyme molecules linked to biotin can be coupled, thereby greatly improving sensitivity;
4 Biotin and avidin have strong affinity, so once they are combined, they are extremely stable, and are not affected by the incubation and multiple washing in the ELISA method, and the binding reaction time is shorter than the time required for the antigen-antibody reaction;
5 The results are highly specific, and the non-specific coloration or staining background is extremely low;
6 The first antibody has a high dilution and can save antibodies.
principle:
When biotin or avidin is combined with an antibody molecule or a label, it does not affect the affinity of the former or the characteristics of the latter. Not all of the four active sites of the avidin molecule bind to the biotin residue attached to the antibody molecule, and the remaining free site is still a receptor for another biotinylated protein. These characteristics are the basis and principle of BAS immunolabeling technology.
BAS basic detection methods can be divided into two categories: one is centered with free avidin, and the biotinylated macromolecular reaction system and labeled biotin are respectively linked, called BAB method or bridged avidin biotin method (bridged avidinbiotintechnique) , BRAB), the improved method is called (avidinbiotincomplex, ABC) method. Another class of biotinylated macromolecular reaction systems linked by labeled avidin is called the BA method or the labeled avidin and biotin method (LAB).
1. The BA method is also known as the LBA method. The specific antibody is biotinylated, and the enzyme molecule is labeled on the avidin molecule. After the biotinylated antibody binds to the test antigen, the enzyme molecule is linked to the antigen molecule by the high affinity of the BAS, and the enzymatic reaction is carried out. The antigen can be detected.
2. The BAB method is also known as the BRAB method. The specific antibody is biotinylated, the enzyme molecule is labeled on the biotin, and then the free avidin bridge is used to integrate the test antigen, the biotinylated antibody and the enzyme-labeled biotin.
3. The principle of ABC method is basically the same as that of BAB method, except that avidin and enzyme biotin need to form ABC complex in a certain proportion. This network structure combines a large number of enzyme molecules, when avidin has not been labeled by enzymes. When the element is saturated, the biotinylated antibody can bind to it. Three basic assays for BAS ELISA.
In the above method, if the second antibody is biotinylated, it is an indirect method. Since the indirect method increases the primary antigen-antibody reaction, it is more sensitive than the direct method and has a wider range of applications. In recent years, many scholars have improved the above basic methods, and some people use avidin antibody or protein A as a bridge. In addition, BA ELISA competition and inhibition methods have been developed.