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An immunonephelometric procedure for determination of apolipoprotein B in human serum [Clin. Chem, 25, 221-226 (1979)] was modified by using the detergent octadecenylamine-polyoxyethylene, which may bind selectively to the phospholipid moiety of lipoproteins. Preincubation of hyperlipemic serum samples with lipases could then be omitted.

The previously described procedure of direct determination of LDL-cholesterol after selective extraction of VLDL and HDL with polycations and a polycation-exchange resin has been improved by using dodecoylated poly(ethyleneimine) instead of poly(ethyleneimine). The acyl derivative has a higher affinity for lipoproteins. Disc-electrophoretic patterns of lipoproteins separated by this procedure correspond to patterns of apolipoproteins in VLDL and HDL, a finding that corroborates the conclusion that the mechanism of the selective binding of certain lipoproteins to lipophilic water-soluble polycations is based on hydrophobic interaction between the polymer and lipids located at the surface of lipoprotein particles. The procedure offers a simple possibility for direct LDL-cholesterol and LDL-triglyceride determination and for the isolation of VLDL and HDL. The mechanism of the reaction has a biochemical correlate in the action of phospholipase A2 on phospholipids.

The previously described procedure of direct determination of LDL-cholesterol after selective extraction of VLDL and HDL with polycations and a polycation-exchange resin has been improved by using dodecoylated poly(ethyleneimine) instead of poly(ethyleneimine). The acyl derivative has a higher affinity for lipoproteins. Disc-electrophoretic patterns of lipoproteins separated by this procedure correspond to patterns of apolipoproteins in VLDL and HDL, a finding that corroborates the conclusion that the mechanism of the selective binding of certain lipoproteins to lipophilic water-soluble polycations is based on hydrophobic interaction between the polymer and lipids located at the surface of lipoprotein particles. The procedure offers a simple possibility for direct LDL-cholesterol and LDL-triglyceride determination and for the isolation of VLDL and HDL. The mechanism of the reaction has a biochemical correlate in the action of phospholipase A2 on phospholipids.

We studied the development of light scattering in the reaction between anti-apolipoprotein B and apolipoprotein B in intact very-low-density lipoproteins (I) and low-density lipoproteins (II) as well as in lipoproteins treated with lipases, and found considerable differences in the kinetics of the immunoreaction for the two lipoprotein classes. Pre-incubation with triglyceride lipase and cholesterol esterase caused a decrease of final light scattering in I but only minimal changes in the reaction with II. Non-ionic detergent not only decreased the original light scattering in hyperlipemic serum samples, but also accelerated the immunoreaction. Under standardized conditions, results of quantitative nephelometry correlated highly significantly with quantitative determination of apolipoprotein B by radial immunodiffusion, both for normolipemic and hyperlipoproteinemic serum samples. The nonspecific light scattering caused by neutral lipids in intact lipoproteins could be minimized when samples were pre-incubated with lipolytic enzymes.

We compared direct determination of beta-lipoprotein cholesterol after selective extraction of very-low-density and high-density lipoproteins from serum with poly(ethyleneimine) and a cation-exchange resin with the classical quantitation after lipoprotein fractionation with the ultracentrifuge. At beta-lipoprotein cholesterol concentrations between 1.50 and 5.00 g/liter the correlation is linear (r = 0.95). The precision for the extraction procedure is as good (CV 2.4-2.8%) as for the quantitation by ultracentrifugation (CV 3.2-6.0%). From solutions of isolated lipoproteins, very-low-density lipoproteins are 93% extracted and high-density lipoproteins 60%, but low-density-lipoproteins only 5%. The molecular mechanism of the extraction is supposed to be due to both hydrophobic interaction of long-chain fatty acid residues and ionic interaction of phospholipids located at the surface of very low-density and high-density lipoproteins and the lipophilic polycation.