检验医学 ›› 2013, Vol. 28 ›› Issue (3): 225-228.DOI: 10.3969/j.issn.1673-8640.2013.03.015

• 技术研究与评价.论著 • 上一篇    下一篇

基于纳米技术的芯片电泳快速检测脂蛋白及其亚型的研究

汪骅1,韩崇旭1,王惠民2,金庆辉3,王大新1,曹丽1   

  1. 1. 苏北人民医院临床医学检测中心，江苏 扬州 225001；2. 南通大学附属医院检验医学中心，江苏 南通 226001；3. 中国科学院上海微系统与信息技术研究所，上海 205001
  • 收稿日期:2012-04-11 修回日期:2012-04-27 出版日期:2013-03-30 发布日期:2013-03-14
  • 通讯作者: 董兰梅，联系电话：0514-87950300。
  • 作者简介:汪骅，男，1975年生，硕士，副主任技师，主要研究方向为微流控技术在临床检验医学中的应用。
  • 基金资助:

    中国国家基础研究发展计划(973计划，2007CB936104)；江苏省社会发展基金资助项目（BE 2010697）； 扬州大学临床医学院资助项目(yzucms 201041)

Research on the lipoprotein and its subclass determination by nanotechnology-based microchip capillary electrophoresis

  1. 1. Clinical Testing Center，Northern Jiangsu People′s Hospital，Jiangsu Yangzhou 225001，China; 2. Laboratory Medicine Center，Affiliated Hospital of Nantong University，Jiangsu Nantong 226001，China; 3. Shanghai Institute of Microsystem and Information Technology，Chinese Academy of Science，Shanghai 225001，China
  • Received:2012-04-11 Revised:2012-04-27 Online:2013-03-30 Published:2013-03-14

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摘要： 目的　建立芯片电泳分离血清脂蛋白及其亚型的方法并初步探讨其临床应用价值。 方法　利用自制的微流控芯片，选择Tricine 缓冲液作为电泳缓冲液，纳米金作为添加剂，分离经硝基苯并噁二唑-C6-酰基鞘胺醇（NBD C6-ceramide）预染的脂蛋白，激光诱导荧光技术检测。探讨不同颗粒大小、不同浓度的纳米金对脂蛋白分离的影响，选择最佳的电泳条件；确定芯片电泳分离血清脂蛋白的线性范围、检测限和重现性；采用芯片电泳分析35例冠心病（CHD）患者与30名健康体检者（对照组）的血清脂蛋白。  结果　在缓冲液中添加20 nmol/L 5 nm纳米金，大而轻低密度脂蛋白（lLDL）、小而密低密度脂蛋白（sdLDL）、极低密度脂蛋白（VLDL）和高密度脂蛋白（HDL）在3 min内实现基线分离；在优化的分离条件下，lLDL、sdLDL、VLDL和 HDL芯片电泳的线性范围[信噪比（S/N）＝3]分别为0.01～0.8、0.04～1.0、0.04～1.0、0.02～0.8 mg/L；检出限分别为5、5、15和 8 μg/L。对健康体检者的血清样本连续测定5次，lLDL、sdLDL、VLDL和HDL的峰面积相对标准偏差（RSD）为分别3.5%、2.2%、4.5%、3.9%。CHD组sdLDL与VLDL水平增高，HDL水平降低，与对照组比较差异均有统计学意义(P＜0.01、P＜0.001)。 结论　基于纳米技术的芯片电泳可快速、简便地测定脂蛋白及其亚型，对评价CHD的危险因素，实现对CHD的早期检测、预防及有效制定CHD防治策略具有重大意义

关键词: 脂蛋白, 纳米金, 芯片电泳, 冠心病

Abstract: Objective　To establish a method for separating serum lipoprotein and its subclass by microchip capillary electrophoresis，and investigate the significance of clinical application.  Methods　The pre-stained lipoproteins with NBD C6-ceramide were separated by Tricine buffers with gold nanoparticles as an additive in quartz chip with laser induced fluorescence detector. The impact of gold nanoparticle with different sizes and concentrations on the separation of lipoprotein was investigated. The best electrophoresis condition was obtained. The detection limit，linearity and reproducibility were investigated. Serum lipoproteins of 30 patients with coronary heart disease (CHD) and 30 healthy subjects (control group) were separated by microchip capillary electrophoresis.  Results　The 20 nmol/L 5nm gold nanoparticles were used as an additive to the buffer in order to obtain the absolute separation of lipoproteins [large and buoyant low density lipoprotein (LDL) (lLDL)，small and dense LDL (sdLDL)，very low-density lipoprotein (VLDL) and high-density lipoprotein(HDL)]. Under optimized conditions，the linear ranges of lLDL，sdLDL，VLDL and HDL were 0.01-0.8，0.04-1.0，0.04-1.0 and 0.02-0.8 mg/L，and their limits of detection were 5，5，15 and 8 μg /L，respectively [signal-to-noise ratio (S/N) ＝ 3]. Relative standard deviation (RSD) values of the peak areas of lLDL，sdLDL，VLDL and HDL were 3.5%，2.2%，4.5% and 3.9%，respectively. The levels of sdLDL and VLDL increased significantly，and HDL decreased more significantly in CHD group than in control group.  Conclusions　The results show that nanotechnology-based microchip capillary electrophoresis is applicable for the rapid and convenient detection of lipoprotein and its subclass，and it has significance for evaluating the analysis of CHD risk factors and the early detection and prevention of CHD.

Key words: Lipoprotein, Gold nanoparticle, Microchip capillary electrophoresis, Coronary heart disease

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