Dupont-Gillain Ch C, Fauroux C M J, Gardner D C J, Leggett G J
Department of Chemistry, University of Manchester Institute of Science and Technology, PO Box 88, Manchester M60 1QD, United Kingdom.
J Biomed Mater Res A. 2003 Nov 1;67(2):548-58. doi: 10.1002/jbm.a.10092.
The adsorption kinetics of human serum albumin (HSA) on CH3- and COOH-terminated self-assembled monolayers (SAMs) has been investigated using radioassays and atomic force microscopy (AFM). On both surfaces, the amount of HSA adsorbed reached a plateau after 30 min. The plateau level was higher on the CH3 compared to the COOH surface. The adhesion force (Fadh), measured using Si3N4 AFM tips in water, decreased with time of contact with the HSA solution on the CH3 surface. This time-dependent change in the adhesiveness of the adsorbed protein is best explained by a change in the conformation or orientation. In contrast, Fadh was independent of the time of contact with the HSA solution on the COOH surface, indicating that once adsorbed, the HSA molecules do not undergo further conformation or orientation changes. The perturbation induced by scanning with the AFM in water on the adsorbed HSA layers was greater on CH3 surfaces than on COOH surfaces, suggesting a weaker protein-substratum interaction on the CH3-terminated SAMs. This was further confirmed by a stronger desorption of HSA following sodium dodecyl sulfate (SDS) treatment on the CH3 surface compared to the COOH surface. Taken together, these data suggest that for COOH SAMs, (1) there is a strong interaction between HSA and the substratum; (2) there is an absence of reorientation with time; and (3) there is a smaller amount of adsorbed protein at 24 h, possibly due to increased but rapid spreading/denaturation of the protein. On the CH3 surface, less deformation of HSA occurs and the molecules maintain a higher mobility at short adsorption times. AFM measurements performed after aging of an adsorbed HSA layer in buffer suggests the role played by HSA in solution in determining the time-dependent conformation and/or orientation changes.
利用放射性测定法和原子力显微镜（AFM）研究了人血清白蛋白（HSA）在甲基（CH3）和羧基（COOH）封端的自组装单分子层（SAMs）上的吸附动力学。在这两种表面上，吸附的HSA量在30分钟后达到平稳状态。与COOH表面相比，CH3表面的平稳吸附水平更高。在水中使用Si3N4 AFM针尖测量的粘附力（Fadh），随着在CH3表面与HSA溶液接触时间的增加而降低。吸附蛋白粘附性的这种时间依赖性变化，最好用构象或取向的变化来解释。相比之下，Fadh与在COOH表面与HSA溶液的接触时间无关，这表明一旦吸附，HSA分子不会发生进一步的构象或取向变化。在水中用AFM扫描对吸附的HSA层引起的扰动，在CH3表面比在COOH表面更大，这表明在CH3封端的SAMs上蛋白质与基质的相互作用较弱。与COOH表面相比，在CH3表面用十二烷基硫酸钠（SDS）处理后HSA的解吸更强，这进一步证实了这一点。综上所述，这些数据表明，对于COOH SAMs，（1）HSA与基质之间存在强相互作用；（2）不存在随时间的重新取向；（3）在24小时时吸附的蛋白量较少，这可能是由于蛋白的扩散/变性增加但速度较快。在CH3表面，HSA发生的变形较小，并且在短吸附时间内分子保持较高的流动性。在缓冲液中对吸附的HSA层进行老化后进行的AFM测量表明，溶液中的HSA在决定时间依赖性构象和/或取向变化方面所起的作用。
