AbstractA modified method of ultrathin‐layer isoelectric focusing is described. The method allows subtyping of phosphoglucomutase even in samples containing considerable amounts of salts and proteins without pretreatment. Wedge‐shaped polyacrylamide or agarose gels varying in thickness from 50 μm (cathode) to 300 μm (anode) are used. If the right pH gradient is chosen the isoenzymes will focus near the cathode where the gel is ultrathin. The sample is applied anodally where the gel is thickest and the capacity therefore high. Thus the effect of salts and proteins on the pH gradient is diminished. Comparative tests have shown that this method is superior to the conventional techniques using gels with a uniform thickness of 1