10196 Phys. Chem. Chem. Phys., 2012, 14, 10196–10206 This journal is c the Owner Societies 2012 Cite this: Phys. Chem. Chem. Phys., 2012, 14, 10196–10206 Mechanism underlying bioinertness of self-assembled monolayers of oligo(ethyleneglycol)-terminated alkanethiols on gold: protein adsorption, platelet adhesion, and surface forcesw Tomohiro Hayashi,* ab Yusaku Tanaka, a Yuki Koide, a Masaru Tanaka c and Masahiko Hara ab Received 18th April 2012, Accepted 15th May 2012 DOI: 10.1039/c2cp41236e The mechanism underlying the bioinertness of the self-assembled monolayers of oligo(ethylene glycol)-terminated alkanethiol (OEG-SAM) was investigated with protein adsorption experiments, platelet adhesion tests, and surface force measurements with an atomic force microscope (AFM). In this work, we performed systematic analysis with SAMs having various terminal groups (–OEG, –OH, –COOH, –NH 2 , and –CH 3 ). The results of the protein adsorption experiment by the quartz crystal microbalance (QCM) method suggested that having one EG unit and the neutrality of total charges of the terminal groups are essential for protein-resistance. In particular, QCM with energy dissipation analyses indicated that proteins absorb onto the OEG-SAM via a very weak interaction compared with other SAMs. Contrary to the protein resistance, at least three EG units as well as the charge neutrality of the SAM are found to be required for anti-platelet adhesion. When the identical SAMs were formed on both AFM probe and substrate, our force measurements revealed that only the OEG-SAMs possessing more than two EG units showed strong repulsion in the range of 4 to 6 nm. In addition, we found that the SAMs with other terminal groups did not exhibit such repulsion. The repulsion between OEG-SAMs was always observed independent of solution conditions [NaCl concentration (between 0 and 1 M) and pH (between 3 and 11)] and was not observed in solution mixed with ethanol, which disrupts the three-dimensional network of the water molecules. We therefore concluded that the repulsion originated from structured interfacial water molecules. Considering the correlation between the above results, we propose that the layer of the structured interfacial water with a thickness of 2 to 3 nm (half of the range of the repulsion observed in the surface force measurements) plays an important role in deterring proteins and platelets from adsorption or adhesion. Introduction Nonfouling (anti-bioadhesion) surfaces have been widely demanded for various biomedical and biosensing applications, and many types of nonfouling surfaces have been reported so far. However, the mechanisms underlying nonfouling behavior have not been fully clarified at a molecular level. 1 In particular, self-assembled monolayers of oligo(ethyleneglycol)-terminated alkanethiol on gold (OEG-SAMs), which was first reported by Prime and Whitesides, 2,3 have provided a model system of protein- and cell-resistant surfaces. OEG-SAMs have been employed to suppress nonspecific adsorption of non-target mole- cules in biosensing and to pattern cells on solid substrates. 4,5 Although several models, which include electrostatic repulsion due to interfacial ions, 6 tightly bound water molecules, 7–9 and stable interfacial water layers, 10–13 have been proposed to explain the nonfouling behavior of OEG-SAMs, the question of why these monolayers, in which thiol molecules are densely packed compared with polymers grafted on surfaces, repel biomolecules and cells has remained unanswered. The main problem in this field is that there has been little systematic work both on the interfacial behavior of water and a Department of Electronic Chemistry, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama, Kanagawa 226-8502, Japan. E-mail: hayashi@echem.titech.ac.jp b Flucto-Order Functions Research Team, Advanced Science Institute, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198, Japan c Department of Biochemical Engineering, Graduate School of Science and Engineering, Yamagata University, Jonan 4-3-16, Yonezawa 992-0038, Japan w Electronic supplementary information (ESI) available: SEM images of self-assembled monolayers after the platelet adhesion experiments, interaction of the EG3–OH SAM (substrate) with NH 2 –SAM (probe) in pure water, interaction of the EG3–OH SAM (substrate) with NH 2 –SAM (probe) in solution at various pHs, and interaction of EG3–OH SAMs with the OH and C8 SAMs in pure water. See DOI: 10.1039/c2cp41236e PCCP Dynamic Article Links www.rsc.org/pccp PAPER