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Immunodiffusion

First developed in the early 1960's by Morris Goodman, among the simplest of all molecular approaches to higher-level taxa phylogenetics are immunodiffusion comparisons of albumins (and some transferrins). Albumins and transferrins are both blood proteins. The principle behind the technique relies on the fact that antibodies produced by the immune system are incredibly sensitive to differences in the structure of molecules they are exposed to. The antibodies of one species will react when exposed to blood serum, containing the proteins, of another. The antibodies that respond against a specific blood protein, such as albumin, are manufactured and collected from an outgroup species. These antibodies, in the form of an antiserum are then exposed to the blood proteins of the species under investigation.

If an organism is only very distantly related to the outgroup species a strong reaction will take place between the blood serum (containing the protein) and the antibodies, and there is very tight binding between the two. The degree of relatedness between the species being tested is reflected by the strength of reaction when exposed to the antiserum. Species more closely related to the outgroup will exhibit a weaker reaction. The experiment is usually conducted on an agar gel with the test blood protein serums arranged in wells around the test antiserum, or some arrangement that will enable comparisons of the test species. One of several different set-ups on the gel may be used, depending on the investigation in question.

The procedure of immunodiffusion is crude, and Sarich (1993) admits that to many scientists it must appear akin to "seeing an engineering student with a slide rule hanging from his belt strolling on a contemporary campus". The response is simple - it works. Although used rather infrequently today, due to the advancement of other molecular methodologies, in the many years that the procedure has been available, immunodiffusion has elucidated many phylogenetic relationships that have later been substantiated by other techniques.