Modification of the bicinchoninic acid protein assay to eliminate lipid interference in determining lipoprotein protein content.
The bicinchoninic acid (BCA) assay method for the determination of protein has been investigated for its utility in measuring the protein content of plasma lipoproteins. Although other methods, principally those based on the method of Lowry et al. (1951, J. Biol. Chem. 193, 265-275) have been extensively used for this purpose, the tolerance of the BCA method to many commonly encountered detergents and buffers offers a definite advantage over the Lowry-based methods. In this study, lipoprotein protein values obtained by the BCA method were compared to a standard modification of the Lowry et al. procedure since this assay forms the basis of much of the relevant literature. The standard BCA assay was found to overestimate the protein content of very low density lipoprotein by approximately 70% and low density lipoprotein by approximately 30%; high density lipoprotein values compared favorably. Overestimations by the BCA assay paralleled the relative phospholipid content of the lipoprotein fractions. This apparent lipid effect was eliminated by the addition of 2% sodium dodecyl sulfate to samples prior to the analysis. In the presence of this detergent, BCA assay measurements for these three lipoprotein fractions were 97, 90, and 98%, respectively, of the reference assay values.
Morton
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Department of Vascular Cell Biology and Atherosclerosis Research, Cleveland Clinic Foundation, Ohio 44195.
Evans
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D006801:Humans; D008074:Lipoproteins / Q000097:blood* / Q000737:chemistry; D010743:Phospholipids / Q000032:analysis / Q000737:chemistry; D011506:Proteins / Q000032:analysis*; D011804:Quinolines / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D013053:Spectrophotometry
D008074:Lipoproteins; D010743:Phospholipids; D011506:Proteins; D011804:Quinolines; D012967:Sodium Dodecyl Sulfate; C047117:bicinchoninic acid