Rheumatoid factor (RF) can also interfere with the capture coating assay for IgM antibodies, leading to false positive reactions. Therefore, the indirect method of neutralizing IgG has recently been favored. The detection of anti-CMV IgGM and anti-Toxoplasma IgM antibodies using these reagents has been successful.
2.2.7 ABS-ELISA kit method
ABS is an abbreviation for the avidin biotin system. Avidin is a glycoprotein with a molecular weight of 60,000. Each molecule consists of four subunits that bind to biotin. Biotin is a small molecule compound with a molecular weight of 244. Derivatives made from chemically-derived hydroxysuccinimide esters can be used to form biotin-labeled products with various types of molecules, such as proteins and sugars. The labeling method is quite simple. The binding of biotin and avidin has strong specificity. The affinity of biotin and avidin is much greater than that of antigen and antibody, and the two are extremely stable once they are combined. Since one avidin can bind to four biotin molecules, the ABS and ELISA kits can be divided into enzyme labeled avidin-biotin (LAB) and bridging avidin-biotin (ABC). Two types of law. Both replace the enzyme-labeled antibody (antigen) in the original human ELISA kit system with a biotin-labeled antibody (or antigen). In LAB, solid biotin is first reacted with unlabeled avidin, and then enzyme-labeled biotin is added to further increase sensitivity. In the early days, avidin was extracted from egg white. This avidin is a basic glycoprotein and has strong adsorption with polystyrene carriers. It can be used in ELISA to increase the background. Streptavidin extracted from Streptomyces does not have this drawback and there is a tendency to replace the former in ELISA kit applications. As the ABS-ELISA kit uses two more reagents than the ordinary ELISA kit, the operation steps are increased.
The detection of IgM antibodies is used in the early diagnosis of infectious diseases. Indirect ELISA kits are generally only suitable for detection of total or IgG antibodies. For example, direct detection of IgM antibodies using antigen-coated indirect methods usually involves the presence of relatively high concentrations of IgG antibodies, which compete for binding to solid-phase antigens and prevent a portion of IgM antibodies from binding to the solid phase. Therefore, if anti-human IgM is used as a secondary antibody to indirectly measure IgM antibodies, the specimen must first be treated with protein A or anti-IgG to remove IgG interference. The capture coating method is often used to determine the IgM of antibodies in clinical tests. The solid phase was first coated with an anti-human IgM antibody to capture IgM (including specific IgM antibodies to the antigen and non-specific IgM antibodies) in the serum samples. The antigen is then added and this antigen binds only to the specific IgM. The enzymes are then labeled with specific antibodies against the antigen. With the substrate, the coloration is positively correlated with the IgM in the specimen. This method is often used for the early diagnosis of viral infections. The detection pattern of hepatitis A virus (HAV) antibody is shown in Figure 2-7.