Simplified Method for the Detection of Apo(a) Isoforms

摘要

Apolipoprotein a, is a high molecular weight glycoproteic component of Lp(a), a molecule associated with coronary arterial disease. Apo(a) exhibits considerable size herterogeneity due to variable repetitions of the carbohydrate-containing structural unit, termed kringle. There are five different kringle froms and 10 different krings 4 types. Apo(a) polymorphism and molecular weight depend on the number of copies of kringle 4 type 2. In this paper we describe a modified 3.75% and 6% discontinuous polyacrylamide gel system and Western-blot technique that shortness the assay time and improves the identification of apo(a) isoforms with a theoretical error of loss than 1 kringle. The assay uses a standard curve prepared with five different recombinant apo(a) molecules, detected up to 50 ng of protein in Lp(a), showed a maximal resolution of 2 kringles and, with the use of third degree polynominal regression analysis, had an error of 0.01275. The inter-assay coefficient of variation was 1.7,2, and 1.4 for the 14 K , 18 K, and 22K phenotypes, whereas the intrassay coefficient of variation was 0.325, 0.18%, and 0.17%, respectively. It is possible that this modified method will diminish the number of putative null alleles so far detected in various studies, but most of all, we are certain that it can be of use in epidemiological studies due to its ease of use, speed, low cost, and enhanced number of samples that can be tested. Abbreviations: Lop(a) = lipoprotein (a); apo(a) = apolipoprotein (a).