The normal isoelectric focusing method has been used to separate various proteins into fractions. The fractions and the parent proteins have been hydrolysed and the amino-acid composition of each has been determined by ion-exchange chromatography. Evidence was obtained that shows that the parent protein is modified by the normal isoelectric focusing procedure; the sulphur-containing amino-acids cystine and methionine were shown to be present partly in the form of cysteic acid and methionine sulphoxides, in a greater proportion than with hydrolysates of proteins isolated by other separation procedures. Prolonged treatment of the protein by the isoelectric focusing procedure adversely affects other amino-acid residues in the molecule also. An improved method has been devised so as to protect the protein isolated by the isoelectric focusing procedure and to prevent the oxidation or modification of the protein, which would otherwise occur during the normal isolation procedure.