title: Enzymatic assessment of cholesterol on electrophoresis gels for estimating HDL size distribution and plasma concentrations of HDL subclasses.
abstract: The aim of this study was to develop an enzymatic cholesterol staining method to determine HDL subclasses in a polyacrylamide gradient gel electrophoresis, which further allows staining by protein in the same electrophoresis lane. HDLs from 120 healthy individuals were separated through nondenaturing PAGE. HDLs were stained for cholesterol using an enzymatic semisolid mixture. Once the gels were unstained, they were stained again for proteins with Coomassie blue. The proportions of HDL subclasses were determined by densitometry. HDL subclasses were transformed to concentrations using as reference HDL-cholesterol plasma levels. This method is comparable in linearity and reproducibility to Coomassie blue staining, although it provides quantitative data. As expected, HDL size distribution shifted toward larger particles when determined by cholesterol as compared with protein. With this method, we observed different proportions of HDL subclasses between men and women as compared with Coomassie blue staining. We described a method to determine HDL size distribution by enzymatic cholesterol staining on polyacrylamide gels. The method allows the quantification of the cholesterol plasma concentration of each HDL subclass with the possibility to further stain the protein in the same sample. The combination of HDL staining by cholesterol and protein on electrophoresis gels provides information that may have clinical relevance.
authors.affiliation: Department of Molecular Biology, Instituto Nacional de Cardiología Ignacio Chávez, Mexico City, Mexico.
authors.forename: Paola
authors.initials: P
authors.lastname: Toledo-Ibelles
authors.forename: Cynthia
authors.initials: C
authors.lastname: García-Sánchez
authors.forename: Nydia
authors.initials: N
authors.lastname: Avila-Vazzini
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Carreón-Torres
authors.forename: Carlos
authors.initials: C
authors.lastname: Posadas-Romero
authors.forename: Gilberto
authors.initials: G
authors.lastname: Vargas-Alarcón
authors.forename: Oscar
authors.initials: O
authors.lastname: Pérez-Méndez
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D008076:Cholesterol, HDL / Q000097:blood* / Q000737:chemistry* / Q000145:classification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D004798:Enzymes / Q000378:metabolism*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D055815:Young Adult
chemical_list: D008076:Cholesterol, HDL; D004798:Enzymes