抄録
A simple procedure is presented to separate serum high density lipoprotein (HDL) into the two subfractions (HDL2 and HDL3) by polyacrylamide gel electrophoresis (PAGE). A 6.75% polyacrylamide gel in 7cm long glass tube and the use of Tris/glycine buffer, pH 8.4, accomplished this separation. The serum lipoproteins were pre-stained with Sudan Black B and electrophoresed for about 90 min at 5mA per gel tube. After completion of electrophoresis, densitometry was performed directly on the tubes containing the gels at 600nm.
The β-lipoprotein was retained on the bottom of the stacking gel. The α-lipoprotein migrated into the separating gel, being separated into the two main components. The fast migrating component of α-lipoprotein was identified as HDL3 and the slow one identified as HDL2 by the electrophoresis of either HDL3 or HDL2 separated by sequential ultracentrifugation. The relative amounts of Sudan Black B stained HDL2 and HDL3 were obtained by integrating the densitometrically traced peak areas of the two components respectively. The HDL cholesterol (HDL-C) concentration of the serum was determined by precipitation (heparin-Ca2+) method. The concentration of serum HDL2-C and HDL3-C can be calculated from the results of these two methods; i. e., the densitometric analysis of polyacrylamide gel and the precipitation method.
The HDL2-C and HDL3-C concentrations measured by this PAGE and precipitation method were well correlated with the values obtained by the ultracentrifugation method. The correlation coefficients for HDL2-C and HDL3-C were 0.91 and 0.90, respectively. Thus, the combination of PAGE and precipitation method for HDL2-C and HDL3-C determination appears to be used as a rapid and reliable screening technique for clinical laboratories.
Both the HDL2-C and HDL3-C concentrations of survivors of cerebral infarction determined by the method described above were lower than those of healthy subjects, and these differences were larger in the HDL2-C concentration than in the HDL3-C one.
