HIGH-SENSITIVITY OF LASER-INDUCED FLUORESCENCE DETECTION IN CAPILLARYGEL-ELECTROPHORESIS FOR ACCURATE APOLIPOPROTEIN-E GENOTYPING

Human apolipoprotein E (apoE) is a product of a polymorphic gene. In t he general population, it shows two major mutations, which lead to the appearance of three common alleles encoding for three protein isoform s. This polymorphism is important in the regulation of lipid metabolis m. Accurate apoE phenotyping or genotyping has become essential in cli nical laboratories, since the epsilon 4 allele has been associated wit h cardiovascular and Alzheimer's diseases. Endonuclease restriction is otyping, followed by slab gel electrophoresis, is a rapid and convenie nt method for the investigation of common apoE genotypes. However duri ng the large-scale apoE genotyping of the STANISLAS cohort, we were co nfronted with a partial lack of sensitivity and resolution power of th is traditional method, which sometimes leads to the misclassification of the genotypes epsilon 2/2 and epsilon 3/2. We have over come this d ifficulty by separating the restriction fragments with capillary gel e lectrophoresis linked to laser-induced fluorescence detection. The bas eline resolution was 2 bp, and the sensitivity limit attainable was si milar to that by radioactive detection. The distinction between the ep silon 3/2 and the epsilon 2/2 genotypes became unequivocal, even when only low amounts of DNA were available for amplification.