Lapin, Norman A.. Chemical and biomolecular functionalization of silicon surfaces for biosensing applications. Retrieved from https://doi.org/doi:10.7282/T3R49QZZ
DescriptionThe reliable functioning of biosensors and bioassays is dependent on the robust attachment of active biomolecules to device substrates, such that the structural integrity, organization and appropriate orientation of these molecules at the surface is maintained. To a significant extent, the underlying surface chemistry and molecular organization that these biomolecules come in contact with and attach to affect the properties of the functional overlayer that they compose. In this work, primarily through the use of infrared spectroscopy, we characterize two main types of biosensor platforms including biotin-streptavidin linkage and surface attachment and covalent attachment of protein to sensor surfaces via amines and sulfhydryls. We further observe the effects of several variations in processing conditions on these platforms including initial atmospheric humidity, use of anhydrous versus aqueous solvents in molecular adsorption and the effect of primary molecular layer stability on the organization and characteristics of subsequently adsorbed biolayers.
With infrared spectroscopy, not only do we identify the formation and breaking of chemical bonds in each of the attachment steps, we also monitor changes in the moieties of each layer with changing environmental conditions. We find that changes in ureido moiety vibrational modes of the biotinylated surface occur near 1250 and 1700 cm-1 dependent on the stability of an underlying layer of aminopropyltriethoxysiloxane, on the type of solvent used in biotinylation itself, and on subsequent protein adsorption to and/or rinsing of the biotinylated surface. In covalent attachment studies, we use small molecules in lieu of protein to characterize amine and sulfhydryl chemical bonding to maleimide-terminated surfaces and using infrared polarization techniques, we find that molecular orientation may be restricted upon covalent attachment. Ellipsometry is used in conjuction with infrared absorption area measurements to determine the relative composition of silane and maleimide prior to attachment of protein.
Additionally, fluorescence measurements of labeled protein are used to quantify protein desorption by surface acoustic wave streaming (SAWS). These measurements are correlated with power dose of SAWS operation. In all, these surface characterization methods are found to successfully monitor chemical and biomolecular layer formation and change under a variety of conditions.