A hydrogel-based versatile screening platform for specific biomolecular recognition in a well plate format |
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Authors: | Beer Meike V Rech Claudia Diederichs Sylvia Hahn Kathrin Bruellhoff Kristina Möller Martin Elling Lothar Groll Jürgen |
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Affiliation: | (1) Department of Functional Materials in Medicine and Dentistry, University of W?rzburg, Pleicherwall 2, 97070 W?rzburg, Germany;(2) Laboratory for Biomaterials, Institute for Biotechnology and Helmholtz Institute for Biomedical Engineering, RWTH Aachen University, Pauwelsstr. 20, 52074 Aachen, Germany;(3) Interactive Materials Research Institute (DWI e.V.) and Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Forckenbeckstr. 50, 52074 Aachen, Germany; |
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Abstract: | Precise determination of biomolecular interactions in high throughput crucially depends on a surface coating technique that allows immobilization of a variety of interaction partners in a non-interacting environment. We present a one-step hydrogel coating system based on isocyanate functional six-arm poly(ethylene oxide)-based star polymers for commercially available 96-well microtiter plates that combines a straightforward and robust coating application with versatile bio-functionalization. This system generates resistance to unspecific protein adsorption and cell adhesion, as demonstrated with fluorescently labeled bovine serum albumin and primary human dermal fibroblasts (HDF), and high specificity for the assessment of biomolecular recognition processes when ligands are immobilized on this surface. One particular advantage is the wide range of biomolecules that can be immobilized and convert the per se inert coating into a specifically interacting surface. We here demonstrate the immobilization and quantification of a broad range of biochemically important ligands, such as peptide sequences GRGDS and GRGDSK-biotin, the broadly applicable coupler molecule biocytin, the protein fibronectin, and the carbohydrates N-acetylglucosamine and N-acetyllactosamine. A simplified protocol for an enzyme-linked immunosorbent assay was established for the detection and quantification of ligands on the coating surface. Cell adhesion on the peptide and protein-modified surfaces was assessed using HDF. All coatings were applied using a one-step preparation technique, including bioactivation, which makes the system suitable for high-throughput screening in a format that is compatible with the most routinely used testing systems. |
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