In an indirect ELISA method the binding of the antigen and antibody is identified by an antibody raised in a second animal. what is the purpose of raising the second antibody in a different animal compared to the other ELISA methods. Under what special cases is this indirect ELISA used
Kamesh, the indirect ELISA principle means that a secondary antibody (AB, which is labeled with peroxidase or whatever) is used. It is obvious that it needs to be from a different species than the first antibody, as one cannot raise an antibody against IgG from the same species, e.g rabbit-anti-rabbit.
Indirect ELISA is the method of choice as primary antibodies usually are not commercially available with a label suitable for detection, like HRP or a fluorescent dye, while providing labeled secondaries is easy and cheap, because there is only a limited number of host animals used for the primary antibodies. Of course, nobody may stop you from labeling your primary antibody with HRP or whatever yourself. It's just not practical, as you'll have to validate and characterize each batch after labeling.
Besides that, when a polyclonal secondary AB is used, there is some chance that more than one molecule will bind to a primary AB, so the signal strength is likely to increase and possibly the sensitivity, too.
I support the comments of Wolfgang and want to add that this assay format is used very frequently and the development is relatively easy. You should test it as one of the first ones. Only if you have specific reasons to avoid this format, you should test other ones. However, you need to have a good antibody for your assay. With a poor antibody, no format will work. And do not forget to include positive and negative controls to identify any problems immediately.
