The plate will require washing with a detergent between each step which can remove the proteins and antibodies not bound. Afterward, the plate is developed via adding some enzymatic substrate which can create the signal demonstrating the amount of antigen in that sample.
As mentioned, the standard signaling method is a color change which is achieved by using a chromogenic reporter and substrates which create an observable colour change. However ELISA biomarkers have developed somewhat and other methods are now also used. For instance some similar techniques now use fluorogenic materials which glow fluorescent, electrochemilumiscent which also illuminate and real time PCR reporters which show quantifiable signals.
There are certain advantages to using these various kinds of reporters. For instance some of them are more sensitive and multiplexing. Newer assays are not technically ELISAs because the name itself and definition suggests the use of enzymes (‘Enzyme Linked’). However they are linked to other reporters but are otherwise the same.
While it is fairly easy to understand, ELISA biomarkers involve a complex process and most pharmaceutical will outsource these tests.
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