Modifications to the technique of two-dimensional immunoelectrophoresis.

In recent years a number of sensitive immunological techniques have been developed to study proteins in biological fluids. The most recently described is that of Minchin Clarke & Freeman (1966) which is a variant of an immunoelectrophoretic method originally described by Ressler (1960) and given the name of crossed immunoelectrophoresis by Laurel1 (1966). It is highly sensitive and by using a combination of gel fractionation and the Minchin Clarke & Freeman (1966) technique, Freeman & Smith (1970) claim that more than sixty immunologically distinct proteins can be recognized. With undiluted serum the present authors can recognize as a routine more than thirty individual proteins. The method consists of an initial electrophoretic separation on a narrow agarose strip; this is then transferred to a second plate, the remainder of which is filled with agarose containing antiserum. The current is next directed at right angles to the initial direction and the partially separated proteins are driven into the bed of antiserum. Each protein appears as a precipitin curve, the area under which is proportional to the concentration of that protein provided the concentration of the antiserum present is constant. Because this technique is so expensive the procedure has been re-examined and a number of modifications have been introduced which make it more suitable for routine use.