The chromogenicity and quantitation of apoB-100 and apoB-48 of human plasma lipoproteins on analytical SDS gel electrophoresis.
ApoB-100 and apoB-48 may be readily resolved in 3.3% sodium dodecyl sulphate-polyacrylamide gels. This study has characterized the relative chromogenicities (staining intensity/micrograms protein) of human apoB-100 and apoB-48 in various lipoprotein classes with Coomassie Brilliant Blue (R250) upon SDS-PAGE. The relation between dye uptake and the mass of each apoB species in any lipoprotein preparation, was linear at least within the concentration range of total apoprotein B which is optimally resolved in these gels (20-50 micrograms total apoprotein B), and was a function of the density of the particular lipoprotein fraction under investigation. There was a constant and characteristic difference between the chromogenicity for apoB-100 and that for apoB-48 as determined from the slopes of their respective chromogenicity curves. The slope of the lines describing staining intensity vs. protein mass for both apoB-100 and apoB-48 decreased as the density of the lipoprotein fraction increased. The slope of the line for apoB-100 was steeper than that for apoB-48 (i.e. chromogenicity apoB-100 greater than apoB-48) in all lipoprotein fractions where both were present. The relationship between the slopes of the lines for apoB-100 and apoB-48 was constant regardless of the density of the lipoprotein fraction. The chromogenicity curves for apoB-100 and for apoB-48 obtained when lipoprotein samples were applied to gels in concentrations conventionally used for this technique (i.e. 20-100 micrograms total apoB/gel) did not extrapolate to the same point on the ordinate, which precludes the use of a simple ratio or "chromogenicity factor" to describe their relative chromogenicities over this concentration range, Hence, a novel approach was developed to determine the relative mass of apoB-100/apoB-48 in lipoprotein samples, based on their staining characteristics in SDS-PAGE.
D053299:Apolipoprotein B-100; D053283:Apolipoprotein B-48; D001055:Apolipoproteins B / Q000032:analysis* / Q000302:isolation & purification; D002863:Chromogenic Compounds / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006801:Humans; D006949:Hyperlipidemias / Q000097:blood; D007700:Kinetics; D008074:Lipoproteins / Q000097:blood*; D012394:Rosaniline Dyes; D012967:Sodium Dodecyl Sulfate
D053299:Apolipoprotein B-100; D053283:Apolipoprotein B-48; D001055:Apolipoproteins B; D002863:Chromogenic Compounds; D008074:Lipoproteins; D012394:Rosaniline Dyes; D012967:Sodium Dodecyl Sulfate; C004692:coomassie Brilliant Blue