Katarzyna Gajos oral presentation (OB4-Fri1-1-3)
TOF-SIMS imaging and chemical surface analysis of biomolecular functionalization of silicon-based immunosensors
1 Jagiellonian University in Krakow, Lojasiewicza 11, 30-348 Krakow, Poland
2 NCSR Demokritos, Aghia Paraksevi, 15210 Athens, Greece
3 AGH University of Science and Technlogy, Mickiewicza 30, 30-059 Krakow, Poland
Miniaturized silicon-based planar waveguides are suited for label free detection of analytes, providing inherently high sensitivity at reduced costs. To take advantage of these features, uniform and high quality biofunctionalization and assay protocols are required for these optical biosensors. Multi-step protocols for immunosensors typically include: surface activation with organo-silanes, immobilization of probe molecules, blocking free sites with non-functional proteins and specific antigen – antibody binding reaction. Biosensor signal can monitor these stages, but it reflects only cumulative surface concentration of all different molecules. In addition, such monitoring is inconvenient for manufacturing purposes, where robotic spotters are used for spatially resolved immobilization of different probe molecules on one chip.
Here, Time-of-Flight Secondary Ion Mass Spectrometry augmented with multivariate data examination (Principal Component Analysis, PCA) is employed to provide step-by-step insight into such multi-step protocols with molecular discrimination. Biofunctionalization and assay protocols applied to Mach-Zehnder interferometers monolithically integrated on silicon chips for immunochemical detection of bovine κ-casein in goat milk  and mycotoxin Ochratoxin A in cereals  are evaluated. TOF-SIMS chemical imaging is employed to determine conditions for uniform probe immobilization. Barely reported to date in-situ chemical inspection of micro-structured on-chip sensors is performed to study changes in composition of different organic molecules well-resolved by different TOF-SIMS signals. Additionally, such subtle changes in surface chemistry varied between different protocols steps are maximized and classified by PCA analysis. This analysis provides also dominant orientations of antibodies (both primary and secondary ones) affinity-bonded during immunoassay as compared to orientations of the same antibodies directly adsorbed on the surface. This complex protocols characterization results in a significant improvement of assay detection sensitivity. This work was supported by the EU funded project “FOODSNIFFER” (FP7-ICT8-318319).
 K. Gajos et al., Applied Surface Science 385, 2016, 529-542.
 K. Gajos et al., Colloids and Surface B: Biointerfaces 150, 2017, 437-444.