Simplified method for the diameter sizing of serum low-density lipoprotein using polyacrylamide gradient gel...

The appearance of small, dense, low-density lipoprotein in serum has been demonstrated to be associated with increased risk of coronary artery disease. The molecular diameter of low-density lipoprotein is usually measured on the basis of mobility differences on polyacrylamide gel electrophoresis. However, since mobility assessed by this method is seriously affected by the increased levels of serum free fatty acids associated with hypertriglyceridemia, we used polyacrylamide gradient gel electrophoresis to eliminate the interference by fatty acids and devised a simple, precise method of polyacrylamide gradient gel electrophoresis to measure the diameter of small, dense, low-density lipoproteins in serum. We used apoferritin and thyroglobulin, which have a molecular diameter of 12.2 nm and 17.0 nm, respectively, and standard low-density lipoprotein particles having a diameter of 25.7 and 27.0 nm as calibrators, estimated by measurement of negative staining of electron microscopy. We also included apoferritin as an internal standard for polyacrylamide gradient gel electrophoresis. The only stain used was Coomassie brilliant blue, and it was used for lipoprotein staining. When we used low-density lipoprotein of 25.73 nm in diameter as a quality control specimen, the coefficient of variation of the size measurements obtained by our method was less than 1.2%. The new method markedly improved the laboratory procedure for measuring the diameter of low-density lipoproteins.