聚二甲基硅氧烷(PDMS)/玻璃微流控芯片电泳快速分离血清高密度脂蛋白亚类及临床应用研究

Application of Polydimethylsiloxane/Glass Microchip for Fast Electrophoretic Separation of Serum High-density Lipoprotein Subclasses

经梯度密度超速离心,高密度脂蛋白(HDL)分为HDL2和HDL3两亚型。HDL2抑制低密度脂蛋白(LDL)氧化功能受损是冠心病(CHD)发生发展的关键因素。因此,通过对HDL亚类进行分离,从而达到预测和诊断CHD的目的。本研究建立了用PDMS/玻璃微流控芯片快速电泳分离HDL亚类的方法。选择N-十二烷基-β-D-麦芽糖苷(DDM)、十二烷基硫酸钠(SDS)和羟丙基纤维素(HPC)共同修饰脂蛋白和泳道表面。在以含0.3 mmol/L SDS的50 mmol/L 3-(N-吗啉代)丙磺酸(MOPS)(pH 8.0)为样品缓冲液,含0.6%HPC的50 mmol/L MOPS(pH 8.0)为分离缓冲液,分离电压为260 V/cm的优化条件下,HDL2和HDL3在4 min内得到基线分离,二者的出峰时间和峰面积的相对标准差(RSD)分别是2.0%和2.7%,2.0%和2.9%,具有较好的重复性。临床标本研究发现,正常人血清标本可分离出HDL2和HDL3双峰,而CHD患者的HDL2峰面积显著减小,甚至消失。PDMS/玻璃微流控芯片分离HDL亚类是一种简单、快速、高效的用于分析CHD危险因子的方法。

On the basis of density by ultracentrifugation,high density lipoprotein（HDL） can be separated into two major subfractions： HDL2 and HDL3.The capacity of HDL2,but not of HDL2,to inhibit the oxidative modification of low density lipoprotein（LDL） is impaired,which is believed to play a central role in coronary heart disease（CHD）,so HDL subclasses separation is necessary for CHD prediction and diagnosis.In this paper,polydimethylsiloxane（PDMS）/glass microchip was used.We chose DDM,SDS and HPC to modify both lipoprotein and the channel surface to reduce lipoprotein adsorption and improve the resolution of lipoprotein separation.Under the optimal conditions,HDL2 and HDL3 were effectively baseline separated with high reproducibility.RSD values of the migration time and peak areas of HDL2 and HDL3 were 2.1% and 2.7%,and 2.0% and 2.9%,respectively.Serum HDL subclasses of healthy ones and patients with CHD were separated by PDMS-based microchip.Two peaks（HDL2 and HDL3） were detected in serum samples of healthy ones while HDL2 fractional peaks almost disappeared in patients′ entire serum samples.These results suggested that PDMS/glass microchip-based HDL subclasses assay was a simple,rapid,and highly efficient technique for CHD risk factors analysis.