Quantitative Analysis of Surface-Immobilized Protein by TOF-SIMS: Effects of Protein Orientation and Trehalose Additive

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• 作者：Young-Pil Kim ; Mi-Young Hong ; Jinmo Kim ; Eunkeu Oh ; Hyun Kyong Shon ; Dae Won Moon ; Hak-Sung Kim ; Tae Geol Lee
• 刊名：Analytical Chemistry
• 出版年：2007
• 出版时间：February 15, 2007
• 年：2007
• 卷：79
• 期：4
• 页码：1377 - 1385
• 全文大小：406K
• 年卷期：v.79,no.4(February 15, 2007)
• ISSN：1520-6882

文摘

We demonstrate the effects of protein orientation andtrehalose on a quantitative analysis of surface-immobilizedproteins by using time-of-flight secondary ion mass spectrometry (TOF-SIMS). As our model protein, streptavidin(SA) was quantitatively immobilized on a solid surface atdifferent configurations by random or oriented immobilization and subsequently treated with trehalose. Theresulting surface was analyzed by using TOF-SIMS andsurface plasmon resonance (SPR) spectroscopy, wherethe secondary ion spectra from SA were compared withthe surface density of the protein. In the case of orientedimmobilization, the ion peak intensities measured by TOF-SIMS were correlated well with the SPR data, regardlessof the presence of trehalose. Alternatively, trehalosesignificantly increased correlation between TOF-SIMS andSPR data for the randomly immobilized SA. It is likely thata trehalose-treated surface is less vulnerable to denaturation, thus leading to a reliable quantification of surface-immobilized proteins by TOF-SIMS. Our results show thatTOF-SIMS can be used for understanding biophysicalstates such as orientation and denaturation of surface-immobilized proteins as well as for quantifying proteinswithin the field of biosensors and biochips.