The interaction of dyes with proteins on paper with special reference to paper electrophoresis.

In attempts to develop quantitative techniques for the analysis of serum proteins separated by electrophoresis on filter paper, many workers have estimated the amount of dye taken up by the different protein fractions after staining. WVhen comparisons have been made with analysis by the classical technique, quantitative differences have been noted (Cremer & Tiselius, 1950; Kunkel & Tiselius, 1951; Schneider, 1951; Grassmann & Hannig, 1952). Cremer & Tiselius used a factor of 1-6 to correct the difference on the grounds that the globulins had a lower binding power for bromophenol blue than had the albumin. This empirical factor was also used by Flynn & de Mayo (1951). Kunkel & Tiselius (1951) published a comparison of albumin and globulin uptake of bromophenol blue using a three-point plot. Grassmann & Hannig (1952) and Grassmann, Hannig & Knedel (1951), describing a procedure for the quantitative estimation of serum proteins using 'Amidoschwarz 1OB' (naphthalein black 12B 200 of Imperial Chemical Industries Ltd.), pointed out the difference between albumin and y-globulin in binding this dye. Martin (1952) stressed the gross differences in dye-binding capacity that existed between the various serum proteins and criticized the use of empirical correction factors derived from normal serawhen studying abnormal sera. Slater & Kunkel (1953), though they give no data, clearly share this doubt about existing quantitative procedures, for they make no attempts at quantitative measurement of individual proteins in a study of urinary and serum proteins, and in their discussion write: 'it appears likely that quantitative procedures applied to filter-paper electrophoresis may be subject to considerable error....'. The increasing use of ffiter-paper electrophoresis for separation of serum proteins, together with the very clearly expressed doubts by workers about the validity of the quantitative results, suggested that