Simplified isoelectric focusing/immunoblotting determination of apoprotein E phenotype.

摘要: We developed a rapid, accurate method for phenotyping apoprotein E that can be used large-scale population studies. In this method, adapted from the of Kamboh et al. (J Lipid Res 1988;29:1535-43), 10-microL plasma samples are incubated with dithiothreitol and Tween-20 15 min then applied to 5% polyacrylamide gels containing ampholyte (pH 4.5-8) urea (3 mol/L). After 2 h isoelectric focusing, bands made visible by immunoblotting. Utilizing whole plasma, does not require time-consuming ultracentrifugation, delipidation samples, or dialysis. Small amounts required, electrofocusing time is short, as many 160 processed per day. Identification phenotype easily accomplished noting location number protein instead their intensity. Because identification affected sialylation, neuraminidase treatment necessary. Agreement in 301 individuals blinded duplicates was 96%, there 98% concordance results 431 had undergone genetic typing. This thus well suited