Isoelectric Focusing Technology for Mapping Post-Translationally Modified Antigens



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Certain substances present in the body, called antigens, are indicative of disease and its progression or remission. Once we know more about antigens a simple blood test could be used to predict disease and measure the efficacy of treatment. A protein-type antigen may undergo various modifications to its initial molecular structure. These modifications, called post-translational modifications (PTMs), often affect the function of the protein. In the interest of understanding the significance of an antigen’s PTMs, we sought to develop a method by which to sort a pure sample of a particular antigens by its PTMs and measure the reactivity of these PTM subsets to a sample from diseased and control patients. A rectangular agarose-based gel was formed and a test protein (Bovine Serum Albumin) was applied to the gel in various concentrations as a test run in an electrophoresis system. An electric current was applied to the gel in order to perform an isoelectric focusing, which separates the protein by charge. The gel is then stained and imaged using specialized equipment. The initial test run was successful and indicated the concentration at which the protein sample should be applied for a clear reading during the imaging process. The next step in the process is to apply a novel antigen protein to the gel, run the isoelectric focusing, incubate the gel in patient sample, and then analyze the results. We believe this will give us information regarding the relative significance of the PTM to the antigen.