About Immunoelecrophoresis

An immunochemistry technique called immunoelecrophoresis makes use of the precipitation process that happens when antigens and antibodies interact in the zone of equivalence. Swedish scientist Arne Tiselius first described the technique in 1930 and utilized it successfully to isolate serum proteins in 1937. At a pH of 8.6, he separated albumin, beta globulin, and gamma globulin (Moticka 360). He received a Nobel Prize in physiology in 1948 after discovering that gamma globulin possesses antibodies that migrate toward the anode in electric field. In 1946 Jacquis Oudin demonstrated that when antigen and antibody are mixed and incubated, a precipited formed at layer of interception between the two. In 1948, Orjan Ouchterlony modified the technique to observe same reaction but on a semisolid gel where diphtheria toxin was introduced in one well and diphtheria antibody in a second well. In 48hours incubation, precipitin lines were formed and distances could be varied by changing concentrations of either as published in 1949. Immunoelectrophoresis proper was first described by Pierre Graber and Curtis William in 1953 when they combined the separation tecquique in electrophoresis with gel diffusion and has since been devised by other scientists such as Alain Bussard who came up with counter electrophoresis in 1959; Curl Bertil devised crossed electrophoresis and rocket electrophoresis (Moticka 361).

Immunoelecrophoresis technique is founded on the ideologies of electrophoresis and immunodiffusion. During the process electrophoresed antigens based on charge migration react with specific antibodies invariably called antiserum resulting in precipitin bands. The bands form in zone of equivalence and form due to cross linkages between sites of fit in the lattice. The electrophoresis is achieved when molecules are introduced to an electric field resulting in acquisition of charge hence movement to appropriate electrode (United States Pharmacopeia: The National Formulary 720). Negatively charged particles move to the Cathode while positively charge ions move to the anode. This mobility is affected by size of molecules being separated with heavy and large particles being slow. Concentration of gel also determines how fast motion would be. Standard concentration of agarose gel has to be used for uniformity of results and reproducibility. The strength of voltage applied also determines the kinetic energy for movement of molecules. Optimal pH of medium used is critical determinant of complete separation and hence buffer will affect electrophoresis. In the immunodiffusion phase where both the soluble antigens resolved from electrophoresis and antibodies migrate distance between the two components and their concentration determine the speed in which the process will occur (Arstila 8). The procedure can therefore be summarized as follows as decribed by Moticka p362:

Gel with wells alternated with strip is prepared

Antigens are resolved to unit components via electrophoresis using charge

Troughs are sliced into agarose agar gel parallel to electric field

Antibody is introduced into the troughs

Antiserum and antibody are subjected to immuniodiffusion

Arcs of precipitin form at equivalence zone

Precipitin line is indication of formed antibody-antibody complex

The future of immunoelectrophoresis is vested in its modification and application. The method would most likely be widely adopted in detection of serum proteins. This detection is important in characterizing the isotypes of immunoglobulins that aid in diagnosis of certain immunological diseases. Overproduction of proteins is also possible. Antibodies may recognize multi-epitopes hence polyclonal or may be specific and selective to a single epitope hence eliminating possibility of cross reaction. The future would utilize electrophoresis to come up with monoclonal antibodies to destroy harmful antigens implicated in various immunological diseases. In vitro fertilization techniques that would stimulate oocytes for enhancing fertility will also employ immunoelectrophoresis (Feichtinger and Kemeteri 69).



Works Cited

Arstila. "Chapter 2 Theory of Electrophoresis and Immunoelectrophoresis." Immunoelectrophoresis, 3 Apr. 2015, pp. 7-10, doi.org/10.1159/000397987.

Feichtinger, W., and P. Kemeter. Future Aspects in Human in Vitro Fertilization. Springer Berlin Heidelberg, 1987.

Moticka, E. J. A Historical Perspective on Evidence-Based Immunology. Elsevier Science, 2015.

The United States Pharmacopeia: The National Formulary. United States Pharmacopeial Convention, 2006.