用^(125)I测定低水平P-选择素平面位点密度新方法  

A new method of measuring P-selectin site density on planar surface based on ^(125)I labeling

作　　者：凌颖琛[1] 李趣欢[1] 黄冰[1] 张金赫[2] 方颖[1] 

机构地区：[1]华南理工大学生物科学与工程学院,广东广州510006 [2]广州军区广州总医院核医学科,广东广州510010

出　　处：《实验技术与管理》2014年第6期36-40,共5页Experimental Technology and Management

基　　金：国家自然科学基金项目(11272125);中国博士后科学基金项目(2013M542180)

摘　　要：建立了一种基于125I放射性标记并与Infinia Hawkeye 4ECT检测相结合的检测蛋白分子位点密度的新方法。实验结果显示,P-选择素的位点密度与吸附浓度呈线性关系,且P-选择素在玻璃平面上的吸附能力高于在聚苯乙烯平面上的吸附。在相同P-选择素浓度下,对比标记P-选择素和P-选择素单克隆抗体9E1的放射性强度,结果发现,物理吸附作用下大概只有10%的P-选择素是头部向上分布的。对比传统方法,新方法更为快速有效,且应用范围更广,新方法所测定的位点密度结果可以与流动腔实验相结合,从而获得一系列二维分子动力学参数。Measuring the P-selectin site density on planar substrate is the primary event in understanding the mechanism of P-selectin-induced cell adhesion on molecular level through the flow chamber experiment . However ,the sensibility of fluorescence labeling fails to detect the small amount of P-selectin ;and the traditional gamma counter cannot detect the radiation on planar surface although the sensibility of 125 I labeling is enough .Thus ,a new method to detect the density of 125 I labeled protein on planar surface is established by using Infinia Hawkeye 4 ECT . The results illustrate that the P-selectin site density is linear with their concentration ,and the glass planar surface shows better adsorption than polystyrene for P-selectin .For the same P-selectin concentration ,the difference in radiation intensity between direct P-selectin with 125 I and its antibody 9E1 with 125 I label bound on P-selectin suggests that only about 10% coated P-selectins are in its head up position .Comparing with the traditional method ,this method is more efficient and can be applied more broadly .Finally ,the results measured by using this new method can be combined with the flow chamber experiment to obtain a series of 2D molecular dynamic parameters .

关 键 词：蛋白分子 P-选择素 位点密度 碘125标记 SPECT 

分 类 号：Q51[生物学—生物化学]