Four methods for determining total protein compared by using purified protein fractions from human serum

The proportional bias of four methods frequently used for determining low concentrations of protein was evaluated with human serum protein fractions (Cohn Fractions II, III, IV, and V). Each fraction was assigned a protein concentration value as determined by the biuret method, calibrated with purified human serum albumin monomer. All four methods (Coomassie Brilliant Blue dye-binding, the method of Lowry et al., ultraviolet absorption, and immunonephelometry gave acceptable results for Fraction V (albumin). The ultraviolet absorption and the Lowry methods overestimated the three globulin fractions (II, III, and IV), whereas the other two methods underestimated these fractions. The method of Lowry et al. gave the least proportional bias for the globulin fractions.