膜片电极支撑双层脂膜的构建和寡核苷酸探针的固定  

CONSTITUTION OF BILAYER LIPID MEMBRANES ON PATCH-CLAMP PIPETTE SUPPORT AND FIXATION OF OLIGONUCLEOTIDE PROBE

作　　者：刘楠[1] 周焕英[1] 高志贤[1] 岳明祥[1] 陈伍忠[1] 安改红 

机构地区：[1]军事医学科学院卫生学环境医学研究所 [2]63600部队后勤部

出　　处：《解放军预防医学杂志》2006年第4期243-246,共4页Journal of Preventive Medicine of Chinese People's Liberation Army

基　　金：国家自然科学基金项目课题(No.30300291);天津市自然科学基金项目课题(No.023609011)

摘　　要：目的对双层脂膜（Bilayer lipid membranes，BLMs）在膜片电极上的构建和十二烷基修饰的寡核苷酸（Oligonucleotide）探针在BLMs上的固定技术进行研究。方法利用膜片钳放大器系统对细胞膜上离子通道跨膜电流变化极其灵敏的特点，构建一种新型BLMs。调节膜片钳放大器。施加一系列电压后对不同浓度的寡核苷酸探针在BLMs上进行固定。结果双层脂膜在膜片电极上稳定构建，成膜完全稳定之后，拟合直线方程为：I（pA）=7．2153＋0．0054V（mV），r=0．9908（P〈0．01），由短杆菌肽法确认了膜的形成。寡核苷酸探针质量浓度在0～273．65ng·mL^-1范围变化时，与跨膜电流呈线性关系，拟合直线方程为：I（pA）：5．4853＋2．9382C（ng·mL^-1），r=0．9962。施加0～180mV阶跃电压于固定有寡核苷酸探针的BINs上，拟合直线方程为：I（pA）=9．3954＋15．2364V（mV），r=0．9977（P〈0．01）。结论膜片电极支撑双层脂膜成功构建并确证，寡核苷酸探针可以在膜片电极支撑双层脂膜上固定，对施加于膜上的一系列电压响应迅速，灵敏度高，稳定性和重复性良好，为构建以膜片电极支撑BLMs的生物传感器监测环境污染物奠定了良好的基础。Objective To study the constitution of bilayer lipid membranes（BLMs）, on patch - clamp electrode and the fixation of dihexyl （C12） - modified oligonucleotide（single- strand DNA, ssDNA） on BLMs. Methods One kind of novel BLMs was fabricated by using patch - clamp pipette technology characterized by sensitive changes of ionic current through cellular membranes, and confirmed by Gramicidin method. Adjusting the amplifier of patch - clamp, electrochemical studies were focused on fixation on BLMs with different concentrations of ssDNA probe by imposing a series of voltages. Results BLMs were stably constituted. After lull stabilization, the fitting straight line was I （pA） = 7. 2153 ＋ 0. 0054V （mV）, r = 0. 9908（ P 〈 0. 01 ）. Ionic current was linearly related to different ssDNA＇ s concentrations from 0 to 273.65 ng ·mL^-1 with the fitting straight line equation： I（pA） = 5.4853 ＋ 2.9382C（ng·mL^-1 ）, r = 0.9962（ P 〈 0.01）. Imposing 0 - 180 mV step voltages on BLMs fixed by ssDNA probe, the fitting straight line equation was I（pA） = 9.3954 ＋ 15. 2364V （ mV ）, r = 0. 9977 （ P 〈 0. 01 ）. Conclusion Patch - clamp pipette supported - BLMs are successfully constituted and confirmed, and ssDNA probe is able to fix on them, which are of rapid response, high sensitivity, favorable stability and repeatability by imposing a series of voltages. Based on these results, a biosensor with patch - clamp pipette - supported BLMs for detection of environmental pollutants may be established.

关 键 词：双层类脂膜 膜片电极 寡核苷酸 

分 类 号：R318[医药卫生—生物医学工程]