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Neutralized Chimeric Avidin Binding at a Reference Biosensor Surface

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Analytical Science Division, National Physical Laboratory, Teddington, UK
Applied Physics Division, Chalmers University of Technology, SE-412 96 Gothenburg, Sweden
§ BioMediTech, University of Tampere and Fimlab Laboratories, FI-33520 Tampere, Finland
*E-mail [email protected]; Tel +44(20)89436193 (A.G.S.).
Cite this: Langmuir 2015, 31, 6, 1921–1930
Publication Date (Web):January 27, 2015
https://doi.org/10.1021/la503213f
Copyright © 2015 American Chemical Society

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    Abstract

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    We describe the development of a reference biosensor surface, based upon a binary mixture of oligo-ethylene glycol thiols, one of which has biotin at the terminus, adsorbed onto gold as self-assembled monolayers (SAMs). These surfaces were analyzed in detail by X-ray photoelectron spectroscopy (XPS) and secondary ion mass spectrometry (SIMS) to establish the relationship between the thiol solution composition and the surface composition and structure. We report the use of argon cluster primary ions for the analysis of PEG-thiols, establishing that the different thiols are intimately mixed and that SIMS may be used to measure surface composition of thiol SAMs on gold with a detection limit better than 1% fractional coverage. The adsorption of neutralized chimeric avidin to these surfaces was measured simultaneously using ellipsometry and QCM-D. Comparison of the two measurements demonstrates the expected nonlinearity of the frequency response of the QCM but also reveals a strong variation in the dissipation signal that correlates with the surface density of biotin. These variations are most likely due to the difference in mechanical response of neutralized chimeric avidin bound by just one biotin moiety at low biotin density and two biotin moieties at high density. The transition between the two modes of binding occurs when the average spacing of biotin ligands approaches the diameter of the avidin molecule.

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    Details of the preparation and characterization of the NCAvd protein; XPS results and a general method for measuring organic film thicknesses on gold using XPS; and a justification for the use of only one QCM-D overtone in the paper. This material is available free of charge via the Internet at http://pubs.acs.org.

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