title: Apolipoprotein E haplotyping by denaturing high-performance liquid chromatography.
abstract: The apolipoprotein E ( APOE ) gene in humans contains two single-base polymorphisms in exon 4, which result in three common alleles, conventionally named epsilon2 , epsilon3 and epsilon4 . Numerous studies have shown an important association between the epsilon4 variant and an increased risk of Alzheimer's disease; other data suggest a possible linkage of APOE genetic heterogeneity with lipid profile and an increased risk of atherothrombotic stroke and coronary heart disease. APOE genotyping is therefore an increasingly common assay in laboratory medicine. The most widely used technique for APOE genotyping is based upon restriction isotyping, i.e., amplification of the fragment of exon 4 containing the most common sequence variations, followed by enzymatic digestion of the amplicon and fragments analysis by gel electrophoresis. We developed a novel, reliable and fast method that exploits the sensitivity and specificity of denaturing high-performance liquid chromatography in detecting single-nucleotide polymorphisms. We show that, in most cases, with a single chromatographic separation it is possible to correctly identify the six different APOE allelic patterns, without any manipulation after the polymerase chain reaction amplification. When compared to restriction isotyping, our method is much faster, less labor intensive and similarly inexpensive.
authors.affiliation: Section of Chemistry, Faculty of Medicine, University of Brescia, Italy.
authors.forename: Maura
authors.initials: M
authors.lastname: Poli
authors.forename: Luisa Benerini
authors.initials: LB
authors.lastname: Gatta
authors.forename: Roberto
authors.initials: R
authors.lastname: Dominici
authors.forename: Carlo
authors.initials: C
authors.lastname: Lovati
authors.forename: Claudio
authors.initials: C
authors.lastname: Mariani
authors.forename: Alberto
authors.initials: A
authors.lastname: Albertini
authors.forename: Dario
authors.initials: D
authors.lastname: Finazzi
mesh_terms: D000483:Alleles; D001057:Apolipoproteins E / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid / Q000379:methods / Q000706:statistics & numerical data; D004247:DNA / Q000235:genetics; D014644:Genetic Variation; D006239:Haplotypes; D006579:Heterozygote; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009691:Nucleic Acid Denaturation; D020641:Polymorphism, Single Nucleotide; D012680:Sensitivity and Specificity
chemical_list: D001057:Apolipoproteins E; D004247:DNA