Wagner M S, Shen M, Horbett T A, Castner David G
National ESCA and Surface Analysis Center for Biomedical Problems, Department of Chemical Engineering, Seattle, Washington 98195-1750, USA.
J Biomed Mater Res A. 2003 Jan 1;64(1):1-11. doi: 10.1002/jbm.a.10263.
Time of flight secondary ion mass spectrometry (ToF-SIMS) is an ideal technique for the analysis of adsorbed protein films because of its surface sensitivity and chemical specificity. In this study, we examined ToF-SIMS with the multivariate calibration method partial least squares regression (PLSR) for the determination of the relative abundance of the components in binary protein films adsorbed onto mica, PTFE, and heptyl amine plasma polymer substrates. These results have been compared with independently measured 125I-radiolabeled protein adsorption experiments. By applying PLSR to the ToF-SIMS data, the relative abundance of the components in the binary adsorbed protein films was quantified, and the agreement between the ToF-SIMS and 125I-radiolabeling data was measured by the root mean square prediction error (RMSPE). Differences in protein quantification by PLSR and 125I-radiolabeling ranged from 5 to 25 mass % RMSPE and were highly dependent on the structure of the adsorbed protein film, the substrate surface chemistry and morphology, and the number of latent variables retained in the PLSR model. The limit of detection for the minor component in the adsorbed protein film was found to be approximately 10 mass %. This study demonstrates that the combination of ToF-SIMS and multivariate calibration provide complementary information to 125I-radiolabeling about the composition and structure of binary adsorbed protein films.
飞行时间二次离子质谱（ToF-SIMS）因其表面灵敏度和化学特异性，是分析吸附蛋白质膜的理想技术。在本研究中，我们使用多变量校准方法偏最小二乘回归（PLSR）对ToF-SIMS进行了研究，以测定吸附在云母、聚四氟乙烯和庚胺等离子体聚合物基底上的二元蛋白质膜中各组分的相对丰度。这些结果已与独立测量的125I放射性标记蛋白质吸附实验进行了比较。通过将PLSR应用于ToF-SIMS数据，对二元吸附蛋白质膜中各组分的相对丰度进行了定量，并通过均方根预测误差（RMSPE）来衡量ToF-SIMS和125I放射性标记数据之间的一致性。PLSR和125I放射性标记在蛋白质定量方面的差异范围为RMSPE的5%至25%，并且高度依赖于吸附蛋白质膜的结构、基底表面化学和形态以及PLSR模型中保留的潜在变量数量。吸附蛋白质膜中次要组分的检测限约为10质量%。本研究表明，ToF-SIMS和多变量校准的结合为125I放射性标记提供了关于二元吸附蛋白质膜组成和结构的补充信息。
