Schmitz G, Möllers C, Richter V
Institut für Klinische Chemie und Laboratoriumsmedizin, Universität Regensburg, Germany.
Electrophoresis. 1997 Sep;18(10):1807-13. doi: 10.1002/elps.1150181015.
An analytical free flow capillary isotachophoresis (cITP) procedure for the detailed analysis of lipoproteins on commercially available capillary electrophoresis systems has been developed. The technique is based on the specific staining of lipoproteins with the fluorescent lipophilic dye 7-nitrobenz-2-oxa-1,3-diazole (NBD)-ceramide before separation. Prestained lipoprotein samples are applied between leading and terminating buffer and separated into 9 well-characterized subpopulations according to their electrophoretic mobility in the absence of any molecular sieve effect. High density lipoproteins are separated into three major subpopulations: (i) the fast migrating high density lipoprotein (HDL) subpopulation (alpha-HDL, containing mainly apolipoprotein (apo) A-I and phosphatidylcholine, (ii) the subpopulation with intermediate mobility, consisting of particles rich in cholesterol, apo A-II, apo E and C apolipoproteins, and (iii) the slow migrating HDL subpopulation (pre-beta-HDL), containing particles rich in apo A-I, apo A-IV. The majority of HDL-associated lecithin:cholesterol acyltransferase (LCAT) activity is also associated with the last subpopulation. The apo B-containing lipoproteins can be subdivided into three major functional groups. The first represents chylomicron derived particles and large triglyceride-rich very low density lipoproteins (VLDL). The second group consists of small VLDL and intermediate density lipoprotein (IDL) particles, and the third group represents the low density lipoproteins (LDL). Results obtained by the isotachophoretic lipoprotein analysis revealed a good correlation in the range of HDL with routinely used techniques, like lipoprotein electrophoresis, HDL-cholesterol analysis by a precipitation procedure or turbidimetric determination of apo A-I. Similar correlations with other analytical techniques were found for the quantitation of the apo B-containing lipoproteins. Advantages of the isotachophoretic separation compared to zone electrophoresis are the high resolution combined with small sample volumes. Moreover, lipoprotein analysis can be performed directly from whole serum, plasma, lymph and other biological fluids in a short time. With these characteristics analytical capillary isotachophoresis may be a helpful tool for a fast and reliable automated quantitation of lipoprotein subpopulations in the clinical laboratory.
已开发出一种分析性自由流动毛细管等速电泳（cITP）方法，用于在市售毛细管电泳系统上对脂蛋白进行详细分析。该技术基于在分离前用荧光亲脂染料7-硝基苯-2-恶唑-1,3-二氮杂萘（NBD）-神经酰胺对脂蛋白进行特异性染色。将预先染色的脂蛋白样品施加在先导缓冲液和终止缓冲液之间，并根据其在不存在任何分子筛效应的情况下的电泳迁移率分离成9个特征明确的亚群。高密度脂蛋白被分离成三个主要亚群：（i）快速迁移的高密度脂蛋白（HDL）亚群（α-HDL，主要包含载脂蛋白（apo）A-I和磷脂酰胆碱），（ii）迁移率中等的亚群，由富含胆固醇、apo A-II、apo E和C载脂蛋白的颗粒组成，以及（iii）缓慢迁移的HDL亚群（前β-HDL），包含富含apo A-I、apo A-IV的颗粒。大多数与HDL相关的卵磷脂：胆固醇酰基转移酶（LCAT）活性也与最后一个亚群相关。含apo B的脂蛋白可细分为三个主要功能组。第一组代表乳糜微粒衍生颗粒和富含大甘油三酯的极低密度脂蛋白（VLDL）。第二组由小的VLDL和中间密度脂蛋白（IDL）颗粒组成，第三组代表低密度脂蛋白（LDL）。通过等速电泳脂蛋白分析获得的结果显示，在HDL范围内与常规使用的技术（如脂蛋白电泳、通过沉淀法进行的HDL胆固醇分析或比浊法测定apo A-I）具有良好的相关性。对于含apo B的脂蛋白的定量，也发现了与其他分析技术的类似相关性。与区带电泳相比，等速电泳分离的优点是高分辨率与小样品体积相结合。此外，脂蛋白分析可以在短时间内直接从全血、血浆、淋巴和其他生物体液中进行。具有这些特性，分析性毛细管等速电泳可能是临床实验室中快速可靠地自动定量脂蛋白亚群的有用工具。
