Quantitative Label-Free Characterization of Avidin鈥揃iotin Assemblies on Silanized Glass

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• 作者：Li-Jung Chen ; Jeong Hyun Seo ; Michael J. Eller ; Stanislav V. Verkhoturov ; Sunny S. Shah ; Alexander Revzin ; Emile A. Schweikert
• 刊名：Analytical Chemistry
• 出版年：2011
• 出版时间：September 15, 2011
• 年：2011
• 卷：83
• 期：18
• 页码：7173-7178
• 全文大小：914K
• 年卷期：v.83,no.18(September 15, 2011)
• ISSN：1520-6882

文摘

In this study, a time-of-flight secondary ion mass spectrometer TOF-SIMS, operating in the event-by-event bombardment/detection mode was used to characterize avidin鈥揵iotin assemblies on silane-modified glass substrates. SIMS was used to analyze several variants of the biointerface, including avidin physically adsorbed on a monofunctional acryl silane surface and covalently attached on monofunctional (amine terminated) and bifunctional (amine and acryl terminated) silanes. The goal of these studies was to determine density of avidin and biotin layers chemically or physically adsorbed on silanized glass substrate. An individual impact of a C<sub>60sub> projectile used in this study creates a hemispherical crater (10 nm in diameter) and emits large numbers of secondary ions from the same nanovolume. Thus, a single impact enables one to unfold distinct secondary ions that span the thickness of the assembled film. This method was used to monitor the presence of glass, silane, and protein ions and to estimate the thickness and density of the avidin layer. In addition, we employed the double coincidence mass spectrometry approach to identify ions coemitted from a specific stratum of the biointerface. This approach was used to determine density of biotin and avidin immobilization while eliminating interferences from isobaric ions that originated from other constituents on the surface. Overall, novel TOF-SIMS quantitative approaches employed here were useful for examining complex biointerfaces and determining both lateral and in depth composition of the film.