Development and In-Depth Characterization of Bacteria Repellent and Bacteria Adhesive Antibody-Coated Surfaces Using Optical Waveguide Biosensing

Biosensors

Szilvia Ruszkai-Szaniszló¹  Barnabás Szabó¹  Máté Varga¹  Péter Fürjes²  Anita Bányai²  Kinga Dóra Kovács³  Andras Saftics³  Eniko Farkas³  Beatrix Peter³  Sandor Kurunczi³  Robert Tarr³  Robert Horvath³  Tamás Gerecsei³  Inna Szekacs³  Attila Bonyár⁴  Eszter Ostorházi⁵  Balazs Stercz⁵  Judit Domokos⁵  Dóra Szabó⁵ 

[1] 77 Elektronika Ltd., 1116 Budapest, Hungary;Centre for Energy Research, Microsystems Lab, Institute of Technical Physics and Materials Science, 1121 Budapest, Hungary;Centre for Energy Research, Nanobiosensorics Laboratory, Institute of Technical Physics and Materials Science, 1121 Budapest, Hungary;Department of Electronics Technology, Faculty of Electrical Engineering and Informatics, Budapest University of Technology and Economics, 1111 Budapest, Hungary;Institute of Medical Microbiology, Semmelweis University, 1089 Budapest, Hungary;

关键词: bacteria sensing;    bacteria repellent coatings;    affinity layers;    layer structure;    binding kinetics;    Escherichia coli;   

DOI  :  10.3390/bios12020056

来源: DOAJ

【 摘 要 】

Bacteria repellent surfaces and antibody-based coatings for bacterial assays have shown a growing demand in the field of biosensors, and have crucial importance in the design of biomedical devices. However, in-depth investigations and comparisons of possible solutions are still missing. The optical waveguide lightmode spectroscopy (OWLS) technique offers label-free, non-invasive, in situ characterization of protein and bacterial adsorption. Moreover, it has excellent flexibility for testing various surface coatings. Here, we describe an OWLS-based method supporting the development of bacteria repellent surfaces and characterize the layer structures and affinities of different antibody-based coatings for bacterial assays. In order to test nonspecific binding blocking agents against bacteria, OWLS chips were coated with bovine serum albumin (BSA), I-block, PAcrAM-g-(PMOXA, NH₂, Si), (PAcrAM-P) and PLL-g-PEG (PP) (with different coating temperatures), and subsequent Escherichia coli adhesion was monitored. We found that the best performing blocking agents could inhibit bacterial adhesion from samples with bacteria concentrations of up to 10⁷ cells/mL. Various immobilization methods were applied to graft a wide range of selected antibodies onto the biosensor’s surface. Simple physisorption, Mix&Go (AnteoBind) (MG) films, covalently immobilized protein A and avidin–biotin based surface chemistries were all fabricated and tested. The surface adsorbed mass densities of deposited antibodies were determined, and the biosensor;s kinetic data were evaluated to divine the possible orientations of the bacteria-capturing antibodies and determine the rate constants and footprints of the binding events. The development of affinity layers was supported by enzyme-linked immunosorbent assay (ELISA) measurements in order to test the bacteria binding capabilities of the antibodies. The best performance in the biosensor measurements was achieved by employing a polyclonal antibody in combination with protein A-based immobilization and PAcrAM-P blocking of nonspecific binding. Using this setting, a surface sensitivity of 70 cells/mm² was demonstrated.

【 授权许可】

Unknown