毛细管电泳-激光诱导荧光检测装置的研制及应用

目的　自行组建高效毛细管电泳 -激光诱导荧光分析装置。方法　以波长 5 4 3.5 nm He- Ne激光为激发光源 ,滤色片 -单色仪分光 ,光电倍增管检测 ,用计算机采集和处理电泳图谱。结果　优化了毛细管电泳 -激光诱导荧光检测装置的设计和分析参数 ,用该装置获得了高信噪比毛细管电泳谱。对罗丹明类荧光试剂 (Sulforhdam ineB)溶液的检出限为 10 - 9m ol/ L,线性范围为 1× 10 - 6～ 1× 10 - 9mol/ L。采用该装置检测了分枝杆菌 DNA的 PCR扩增产物、酶切产物以及人血清白蛋白。结论　所组建的高效毛细管电泳 -激光诱导荧光分析装置结构合理 ,灵敏度高 ,成本低、光路校准方便 ,分离效能明显高于常规琼脂糖电泳 ,能作为研究生物样品中 DNA片段和蛋白质含量的有效检测手段。

Study on a Sensitive Setup of Capillary Electrophoresis with Laser Induced Fluorescence Detector and Its Application

OBJECTIVE: To develop a setup of capillary electrophoresis with laser induced fluorescence detector for separating and detecting DNA fragments and protein. METHODS: The setup is assembled of He-Ne Laser with 543.5 nm as the excitation light, filter, monochromator, photomultiplier tube and a computer for controlling the system and collecting the electropherograms. RESULTS: The design of the setup and the analytical parameters were optimized. The electropherograms with high Signal-to-Noise were obtained by the laser induced fluorescence detector-capillary electrophoretic setup. For Sulforhdamine B, the detection limit was 10(-9) mol/L, and the linear range from 1 x 10(-6) mol/L to 1 x 10(-9) mol/L. The PCR products, the digested products by Hae III and BstE of mycobacterium DNA, and the human serum albumin were determined by the successful application of this setup. CONCLUSION: The setup is characterized by simple structure, low cost and high sensitivity. Compared with traditional agarose gel electrophoresis, capillary electrophoresis with laser induced fluorescence detector is more remarkable in resolution and detection time. It can be applied as a more effective means for analysis of DNA fragments and protein.