Tobias Ekblad

Poly(ethylene glycol)-Containing Hydrogel Surfaces for Antifouling Applications in Marine and Freshwater Environments

This work describes the fabrication, characterization, and biological evaluation of a thin protein-resistant poly(ethylene glycol) (PEG)-based hydrogel coating for antifouling applications. The coating was fabricated by free-radical polymerization on silanized glass and silicon and on polystyrene-covered silicon and gold. The physicochemical properties of the coating were characterized by infrared spectroscopy, ellipsometry, and contact angle measurements. In particular, the chemical stability of the coating in artificial seawater was evaluated over a six-month period. These measurements indicated that the degradation process was slow under the test conditions chosen, with the coating thickness and composition changing only marginally over the period. The settlement behavior of a broad and diverse group of marine and freshwater fouling organisms was evaluated. The tested organisms were barnacle larvae (Balanus amphitrite), algal zoospores (Ulva linza), diatoms (Navicula perminuta), and three bacteria species (Cobetia marina, Marinobacter hydrocarbonoclasticus, and Pseudomonas fluorescens). The biological results showed that the hydrogel coating exhibited excellent antifouling properties with respect to settlement and removal.

Gradient Hydrogel Matrix for Microarray and Biosensor Applications : An Imaging SPR Study

A biosensor matrix based on UV-initiated graft copolymerized poly(ethylene glycol) methacrylate and 2-hydroxyethyl methacrylate has been studied using imaging surface plasmon resonance (iSPR). By using a photo mask and a programmable shutter to vary the exposure time laterally, a gradient of matrix spots with physical thicknesses ranging from a few to tens of nanometers was generated. To maximize the dynamic range, imaging SPR was employed in wavelength interrogation mode. By finding the minimum in the reflectance spectra from each pixel of an image, SPR wavelength maps were constructed. The shift in SPR wavelength upon biospecific interaction was then measured both as a function of matrix thickness and composition. The performance of the matrix was evaluated in terms of immobilization of human serum albumin, biomolecular interaction with its antibody, and nonspecific binding of human fibrinogen. In addition, a low molecular weight interaction pair based on a synthetic polypeptide and calmodulin was also studied to explore the size selectivity of the hydrogel matrix. Our results show that the gradient matrix exhibits excellent properties for quick evaluation and screening of optimal hydrogel performance. The mixed hydrogel matrices display very low levels of nonspecific binding. It is also evident that the low molecular weight calmodulin is capable of freely diffusing and interacting throughout the entire hydrogel matrix, whereas the much larger albumin and its corresponding antibody, in particular, are partly/completely hindered from penetrating the interior of the matrix. This size-selectivity is attributed to a significant UV-initiated cross-linking or branching of the matrix during fabrication and/or protein mediated multipoint attachment during immobilization.

Lateral control of protein adsorption on charged polymer gradients.

This work describes the fabrication, characterization, and protein adsorption behavior of charged polymer gradients. The thin gradient films were fabricated by a two-step technique using UV-initiated free-radical polymerization in a reactor with a moving shutter. A homogeneous layer of cationic poly(2-aminoethyl methacrylate hydrochloride) was first formed, followed by a layer of oppositely charged poly(2-carboxyethyl acrylate) with a continuously increasing thickness. Adsorption from protein solutions as well as human blood plasma was investigated by imaging surface plasmon resonance and infrared microscopy. The results showed excessive protein adsorption in the areas where one of the polymers dominated the composition, while there was a clear minimum at an intermediate position of the gradient. The charge of the surface was estimated by direct force measurements and found to correlate well with the protein adsorption, showing the lowest net charge in the same area as the protein adsorption minimum. We therefore hypothesize that a combination of the charged polymers, in the right proportions, can result in a protein-resistant surface due to balanced charges.

Resistance of galactoside-terminated alkanethiol self-assembled monolayers to marine fouling organisms

Self-assembled monolayers (SAMs) of galactoside-terminated alkanethiols have protein-resistance properties which can be tuned via the degree of methylation [Langmuir 2005, 21, 2971-2980]. Specifically, a partially methylated compound was more resistant to nonspecific protein adsorption than the hydroxylated or fully methylated counterparts. We investigate whether this also holds true for resistance to the attachment and adhesion of a range of marine species, in order to clarify to what extent resistance to protein adsorption correlates with the more complex adhesion of fouling organisms. The partially methylated galactoside-terminated SAM was further compared to a mixed monolayer of ω-substituted methyl- and hydroxyl-terminated alkanethiols with wetting properties and surface ratio of hydroxyl to methyl groups matching that of the galactoside. The settlement (initial attachment) and adhesion strength of four model marine fouling organisms were investigated, representing both micro- and macrofoulers; two bacteria (Cobetia marina and Marinobacter hydrocarbonoclasticus), barnacle cypris larvae (Balanus amphitrite), and algal zoospores (Ulva linza). The minimum in protein adsorption onto the partially methylated galactoside surface was partly reproduced in the marine fouling assays, providing some support for a relationship between protein resistance and adhesion of marine fouling organisms. The mixed alkanethiol SAM, which was matched in wettability to the partially methylated galactoside SAM, consistently showed higher settlement (initial attachment) of test organisms than the galactoside, implying that both wettability and surface chemistry are insufficient to explain differences in fouling resistance. We suggest that differences in the structure of interfacial water may explain the variation in adhesion to these SAMs.

Saccharide-functionalized alkanethiols for fouling-resistant self-assembled monolayers: synthesis, monolayer properties, and antifouling behavior.

We describe the synthesis of a series of mono-, di-, and trisaccharide-functionalized alkanethiols as well as the formation of fouling-resistant self-assembled monolayers (SAMs) from these. The SAMs were characterized using ellipsometry, wetting measurements, and infrared reflection-absorption spectroscopy (IRAS). We show that the structure of the carbohydrate moiety affects the packing density and that this also alters the alkane chain organization. Upon increasing the size of the sugar moieties (from mono- to di- and trisaccharides), the structural qualities of the monolayers deteriorated with increasing disorder, and for the trisaccharide, slow reorganization dynamics in response to changes in the environmental polarity were observed. The antifouling properties of these SAMs were investigated through protein adsorption experiments from buffer solutions as well as settlement (attachment) tests using two common marine fouling species, zoospores of the green macroalga Ulva linza and cypris larvae of the barnacle Balanus amphitrite. The SAMs showed overall good resistance to fouling by both the proteins and the tested marine organisms. To improve the packing density of the SAMs with bulky headgroups, we employed mixed SAMs where the saccharide-thiols are diluted with a filler molecule having a small 2-hydroxyethyl headgroup. This method also provides a means by which the steric availability of sugar moieties can be varied, which is of interest for specific interaction studies with surface-bound sugars. The results of the surface dilution study and the low nonspecific adsorption onto the SAMs both indicate the feasibility of this approach.

Blood compatibility of photografted hydrogel coatings.

In this work, we have evaluated the haemocompatibility of different surface modifications, intended for biomaterials and bioanalytical applications. Polystyrene slides were coated with thin hydrogel films by self-initiated photografting and photopolymerization (SIPGP) of four different monomers. The hydrogel surface modifications were thoroughly characterized and tested for their protein resistance and ability to resist platelet adhesion and activation of the coagulation system. There was very little protein adsorption from human plasma on the hydrogels prepared from poly(ethylene glycol) methacrylate and 2-hydroxyethyl methacrylate. Platelet adhesion tests performed under both static and flow conditions showed that these coatings also demonstrated very high resistance towards platelet adhesion. A small amount of platelets were found to adhere to hydrogels formed from ethylene glycol methyl ether methacrylate and 2-carboxyethyl methacrylate. The polystyrene substrates themselves facilitated large amounts of platelet adhesion under both static and flow conditions. Utilizing a novel setup for imaging of coagulation, it was confirmed that none of the hydrogel surfaces activated the coagulation system to any great extent. We suggest that this simple fabrication method can be used to produce hydrogel coatings with unusually high blood compatibility, suitable for demanding biomaterials applications.

Novel application of imaging surface plasmon resonance for in situ studies of the surface exploration of marine organisms.

The surface interactions of exploring cyprids of the barnacle Semibalanus balanoides were studied in situ using imaging surface plasmon resonance. It was demonstrated how the deposition of a proteinaceous adhesive could be followed in real time as the cyprids explored and temporarily attached to a surface. Furthermore, the amount of protein left on the surface when the cyprids moved on could be quantified. Clear differences were demonstrated between an oligo(ethyleneglycol) coated surface and a bare gold substrate. It is anticipated that this technique will be a valuable tool in the development of novel surface chemistries that can prevent biofouling.

Swelling of Thin Poly(ethylene glycol)-Containing Hydrogel Films in Water Vapor—A Neutron Reflectivity Study

Hydrogels are widely used in biomedicine and for bioanalytical purposes, normally under wet conditions. For certain applications, processing steps, or process monitoring, hydrogel films are used or treated under ambient conditions, and because they are hygroscopic, it is of interest to investigate how they respond to changes in atmospheric humidity. We have used neutron reflectometry to follow the swelling of thin UV-polymerized hydrogel films in air under different relative humidities (RHs). These polymers were prepared to similar thicknesses on silica and gold substrates, and the chemical similarity between them was verified by infrared spectroscopy. The swelling in response to variations in RH was different for the layers on the two substrate types, reflecting structural changes induced by differences in the UV exposure required to achieve a given polymer thickness, as demonstrated also by differences in the Flory-Huggins interaction parameter, obtained by fitting a Flory-Huggins-type sorption model to the swelling data. Wetting studies show small changes in contact angles with surrounding humidity variations, indicating that structural reorganization at the interface in response to humidity changes is limited.