Characterization of free and tightly bound lipoprotein in intima by thin layer isoelectric focusing

Thin layer isoelectric focusing followed by second dimension quantitative immunoelectrophoresis has been used to characterize the free and tightly bound lipoprotein (lp) fractions in human aortic intima. In 3.3% acrylamide gels plasma low density lipoprotein (LDL) focuses rapidly, but very low density lipoprotein (VLDL) fails to enter the gell and remains at the origin (point of application). Samples of normal intima and lesions were placed directly on the gel and focused for 10--12 h; in normal intima and gelatinous lesions which had not accumulated cholesterol 60--70% of the free LP focused with plasma LDL, but with increasing accumulation of cholesterol the proportion focusing with LDL decreased and the component remaining at the origin increased. In 9 of the 39 samples examined one or more components with pI acid to LDL were present, and in one sample the whole peak focused at a more acid pI. In 4 of 5 lipid-rich centres of gelatinous plaques all the free LP remained at the origin, and the bound LP released by incubation with plasmin remained at the origin in all samples in 3.3% gels although it would focus in 2.6% gels. With antiserum to apo-C this large LP did not behave like VLDL, and it appears to be an aggregated LDL. An increased proportion of aggregated LDL was associated with (a) accumulation of cholesterol, (b) topographical location towards the centre of plaques, and (c) partial destruction of endogenous LP during incubation of fresh samples of tissue.