CLINICAL-CHEMISTRY OF COMMON APOLIPOPROTEIN-E ISOFORMS

Apolipoprotein E plays a central role in clearance of lipoprotein remn ants by serving as a ligand for low-density lipoprotein and apolipopro tein E receptors. Three common alleles (apolipoprotein E(2), E(3) and E(4)) give rise to six phenotypes. Apolipoprotein E(3) is the ancestra l form. Common apolipoprotein E isoforms derive from nucleotide substi tutions in codons 112 and 158. Resulting cysteine-arginine substitutio ns cause differences in: affinities for low-density lipoprotein and ap olipoprotein E receptors, low-density lipoprotein receptor activities, distribution of apolipoprotein E among lipoproteins, low-density lipo protein formation rate, and cholesterol absorption. Accompanying chang es in triglycerides, cholesterol and low-density lipoprotein may promo te atherosclerosis development. Over 90% of patients with familial dys betalipoproteinaemia have apolipoprotein E(2)/E(2). Apolipoprotein E(4 ) may promote atherosclerosis by its low-density lipoprotein raising e ffect. Establishment of apolipoprotein E isoforms may be important for patients with diabetes mellitus and several non-atherosclerotic disea ses. Apolipoprotein E phenotyping exploits differences in isoelectric points. Isoelectric focusing uses gels that contain pH 4-7 ampholytes and urea. Serum is directly applied, or prepurified by delipidation, l ipoprotein precipitation or dialysation. Isoelectric focusing is follo wed by immunofixation/protein staining. Another approach is electro- o r diffusion blotting, followed by protein staining or immunological de tection with anti-apolipoprotein E antibodies and an enzyme-conjugated second antibody. Apolipoprotein E genotyping demonstrates underlying point mutations. Analyses of polymerase chain reaction products are do ne by allele-specific oligonucleotide probes, restriction fragment len gth polymorphism, single-stranded conformational polymorphism, the pri mer-guided nucleotide incorporation assay, or denaturating gradient ge l electrophoresis. Detection with primers that either or not initiate amplification is performed with the amplification refractory mutation system. Disparities between phenotyping and genotyping may derive from isoelectric focusing methods that do not adequately separate apolipop rotein E posttranslational variants, storage artifacts or faint isoele ctric focusing bands.