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4 Types of ELISA (Direct, Indirect, Sandwich, Competitive)

Immunoassays all utilize the bond between an antibody and its specific antigen for them to work. The two main categories of immunoassays are unlabeled and labeled. Labeled immunoassays consist of radio immunoassays and the much more frequently used Enzyme-linked immunosorbent assay or ELISA which we will look at today. There are four different types of ELISA, direct, indirect, sandwich and competitive ELISA. In direct ELISA the target antigen has been bound to the floor of the micro well plate. Then primary monoclonal antibodies specific to the target antigens bind to them. Moreover, these antibodies have been labeled with an enzyme which can interact with a substrate to produce a measurable signal. Indirect ELISA works in the exact same manner but here an additional secondary antibody binds to the primary antibody. This secondary antibody that has been outfitted with a label which produces a signal in the same manner as in the previous case. In a sandwich ELISA, a so-called capture-antibody has been bound to the floor of the well. Then when the sample is added, the target analyte binds to the capture antibody. Then another antibody, also specific to the target analyte binds to it, forming a sort of sandwich structure. This second antibody has been labeled and can thus produce a signal. Finally competitive ELISA works slightly different. Here, the antigen is first mixed with an excess of antibodies specific to it. These then react and form an antigen-antibody complex. Then the mixture is added to a micro-well plate containing antigens specific to the antibodies in the mixture. The result is that only unbound antibodies can bind to these antigens. Labeled secondary antibodies then bind to these primary antibodies and produce an inverse signal, meaning that the more signal we get the less antigens were present in out initial mixture of antigens and antibodies.