Cesar Rodriguez-Emmenegger

Polymer Brushes Showing Non-Fouling in Blood Plasma Challenge the Currently Accepted Design of Protein Resistant Surfaces

Ultra-low-fouling poly[N-(2-hydroxypropyl) methacrylamide] (poly(HPMA)) brushes have been synthesized for the first time. Similar to the so far only ultra-low-fouling surface, poly(carboxybetaine acrylamide), the level of blood plasma fouling was below the detection limit of surface plasmon resonance (SPR, 0.03 ng·cm(-2)) despite being a hydrogen bond donor and displaying a moderate wettability, thus challenging the currently accepted views for the design of antifouling properties. The antifouling properties were preserved even after two years of storage. To demonstrate the potential of poly(HPMA) brushes for the preparation of bioactive ultra-low fouling surfaces a label-free SPR immunosensor for detection of G Streptococcus was prepared.

Controlled Cell Adhesion on Poly(dopamine) Interfaces Photopatterned with Non‐Fouling Brushes

Bioinspired poly(dopamine) (PDA) films are merged with antifouling poly(MeOEGMA) brushes utilizing a nitrile imine-mediated tetrazole-ene cycloaddition (NITEC)-based phototriggered surface encoding protocol. The antifouling brushes were photopatterned on PDA surfaces, leading cells to form confluent layers in the non-irradiated sections, while no adhesion occurred on the brushes resulting in a remarkably precise cell pattern. The presented strategy paves the way for the design of tailor-made patterned cell interfaces.

Complete identification of proteins responsible for human blood plasma fouling on poly(ethylene glycol)-based surfaces.

The resistance of poly(ethylene glycol) (PEG) against protein adsorption is crucial and has been widely utilized in various biomedical applications. In this work, the complete protein composition of biofilms deposited on PEG-based surfaces from human blood plasma (BP) was identified for the first time using nanoLC-MS/MS, a powerful tool in protein analysis. The mass of deposited BP and the number of different proteins contained in the deposits on individual surfaces decreased in the order of self-assembling monolayers of oligo(ethylene glycol) alkanethiolates (SAM) > poly(ethylene glycol) end-grafted onto a SAM > poly(oligo(ethylene glycol) methacrylate) brushes prepared by surface initiated polymerization (poly(OEGMA)). The BP deposit on the poly(OEGMA) surface was composed only of apolipoprotein A-I, apolipoprotein B-100, complement C3, complement C4-A, complement C4-B, histidine-rich glycoprotein, Ig mu chain C region, fibrinogen (Fbg), and serum albumin (HSA). The total resistance of the surface to the Fbg and HSA adsorption from single protein solutions suggested that their deposition from BP was mediated by some of the other proteins. Current theories of protein resistance are not sufficient to explain the observed plasma fouling. The research focused on the identified proteins, and the experimental approach used in this work can provide the basis for the understanding and rational design of plasma-resistant surfaces.

Photo‐Patterning of Non‐Fouling Polymers and Biomolecules on Paper

Functional cellulose substrates with tetrazole moieties are generated to serve as universal platforms for the spatio-temporal immobilization of synthetic ultra-low fouling polymer brushes and protein species via a nitrile imine-mediated tetrazole-ene cycloaddition (NITEC)-based protocol. Poly(carboxybetaine acrylamide) brushes are grafted from initiators photo-patterned by NITEC utilizing single electron transfer living radical polymerization. Streptavidin is photo-immobilized with remarkable efficiency, opening the possibility to generate new materials for biomedical and biosensing applications.

Substrate-Independent Approach for the Generation of Functional Protein Resistant Surfaces

A new route for coating various substrates with antifouling polymer layers was developed. It consisted in deposition of an amino-rich adhesion layer by means of RF magnetron sputtering of Nylon 6,6 followed by the well-controlled, surface-initiated atom transfer radical polymerization of antifouling polymer brushes initiated by bromoisobutyrate covalently attached to amino groups present in the adhesion layer. Polymer brushes of hydroxy- and methoxy-capped oligoethyleneglycol methacrylate and carboxybetaine acrylamide were grafted from bromoisobutyrate initiator attached to a 15 nm thick amino-rich adhesion layer deposited on gold, silicon, polypropylene, and titanium-aluminum-vanadium alloy surfaces. Well-controlled polymerization kinetics made it possible to control the thickness of the brushes at a nanometer scale. Zero fouling from single protein solutions and a reduction of more than 90% in the fouling from blood plasma observed on the uncoated surfaces was achieved. The feasibility of functionalization with bioactive compounds was tested by covalent attachment of streptavidin onto poly(oligoethylene glycol methacrylate) brush and subsequent immobilization of model antibodies and oligonucleotides. The procedure is nondestructive and does not require any chemical preactivation or the presence of reactive groups on the substrate surface. Contrary to current antifouling modifications, the developed coating can be built on various classes of substrates and preserves its antifouling properties even in undiluted blood plasma. The new technique might be used for fabrication of biotechnological and biomedical devices with tailor-made functions that will not be impaired by fouling from ambient biological media.

Poly(HEMA) brushes emerging as a new platform for direct detection of food pathogen in milk samples

Surface plasmon resonance (SPR) biosensors capable of in real time detection of Cronobacter at concentrations down to 10⁶ cells mL⁻¹ in samples of consumer fresh-whole fat milk, powder whole-fat milk preparation, and powder infant formulation were developed for the first time. Antibodies against Cronobacter were covalently attached onto polymer brushes of poly(2-hydroxyethyl methacrylate) (poly(HEMA)) grafted from the SPR chip surface. The lowest detection limit, 10⁴ cells mL⁻¹, was achieved in phosphate buffered saline (pH 7.4) with sensors prepared by covalent immobilization of the same antibodies onto a self assembled monolayer (SAM) of hexa(ethylene glycol) undecanethiol (EG₆). However, when the EG₆ based sensors were challenged with milk samples the non-specific response due to the deposition of non-targeted compounds from the milk samples was much higher than the specific response to Cronobacter hampering the detection in milk. Similar interfering fouling was observed on antifouling polymer brushes of hydroxy-capped oligoethylene glycol methacrylate and even a 10 times higher fouling was observed on the widely used SAM of mixed hydroxy- and carboxy-terminated alkanethiols. Only poly(HEMA) brushes totally suppressed the fouling from milk samples. The robust well-controlled surface initiated atom transfer radical polymerization of HEMA allowed the preparation of highly dense brushes with a minimal thickness so that the capture of antigens by the antibodies immobilized on the brush layer could take place close to the gold SPR surface to provide a stronger optical response while the fouling was still suppressed. A minimum thickness of 19 nm of poly(HEMA) brush layer was necessary to suppress completely non-specific sensor response to fouling from milk.

Surfaces resistant to fouling from biological fluids: towards bioactive surfaces for real applications.

The fouling from four human body fluids - blood plasma, cerebrospinal fluid, urine and saliva - and four animal fluids - foetal bovine and calf sera, egg and milk - relevant to human and veterinary medicine, immunology, biology and diagnostics is assessed on antifouling SAMs and on polymer brushes of oligo(ethylene glycol) methacrylate, 2-hydroxyethyl methacrylate, carboxybetaine acrylamide and N-(2-hydroxypropyl)methacrylamide synthesized via ATRP. While important deposits from the all biofluids are observed on SAMs, a superior resistance is achieved on polymer brushes. Importantly, only poly(CBAA) and poly(HPMA) are capable of resisting the fouling from the most challenging media, blood plasma and eggs.

Non-fouling Hydrogels of 2-Hydroxyethyl Methacrylate and Zwitterionic Carboxybetaine (Meth)acrylamides

Five poly(betaine) brushes were prepared, and their resistance to blood plasma fouling was studied. Two carboxybetaines monomers were copolymerized with 2-hydroxyethyl methacrylate (HEMA) to prepare novel hydrogels. By increasing the content of the zwitterionic comonomer, a 4-fold increase in the water content could be achieved while retaining mechanical properties close to the widely used poly(HEMA) hydrogels. All hydrogels showed an unprecedentedly low fouling from blood plasma. Remarkably, by copolymerization with 10 mol % of carboxybetaine acrylamide, hydrogels fully resistant to blood plasma were prepared.

Biomimetic non-fouling surfaces: extending the concepts

In this study, we propose a substrate-independent biomimetic modification route for the creation of antifouling polymer brushes. This modification route consists of the formation/deposition of a biomimetic polydopamine anchor layer followed by a well-controlled surface-initiated atom transfer radical polymerization of antifouling polymer brushes initiated by 2-bromo-2-methylpropanoyl groups covalently attached to the hydroxyl and amine groups present in the anchor layer. In this way, we synthesized polymer brushes of methoxy- and hydroxy-capped oligoethylene glycol methacrylate, 2-hydroxyethyl methacrylate and carboxybetaine acrylamide. Spectroscopic ellipsometry (SE) indicated well-controlled polymerization kinetics of the brushes, thus the thickness of the ultra-thin films could be precisely tuned at a nanometer scale. The covalent structure and organization of the brushes grown from the polydopamine anchor layer were accessed by infrared reflection-adsorption spectroscopy (IRRAS) while the change in hydrophilicity caused by the presence of the brush was determined by dynamic water contact angle measurements. Surface plasmon resonance as well as ex situ IRRAS and SE measurements were applied to investigate the adsorption of model protein solutions and undiluted human blood plasma to the brushes. The biomimetic brushes completely suppressed the fouling from single protein solutions and reduced the fouling from plasma to less than 3% from the fouling measured on bare gold surfaces. The proposed modification procedure is non-destructive and does not require any chemical pre-activation or the presence of reactive groups on the substrate surface. Contrary to other antifouling modifications the coating can be performed on various classes of substrates and preserves its properties even in undiluted blood plasma. This work offers a promising technology for the facile fabrication of different surface-based biotechnological and biomedical devices able to perform tailor-made functions while resisting the fouling from the complex biological media where they operate.

SET-LRP of N-(2-hydroxypropyl)methacrylamide in H2O

Cu(0) wire-catalyzed SET-LRP of N-(2-hydroxypropyl)methacrylamide (HPMA) initiated with methyl 2-chloropropionate (MCP) in H2O at 50 °C exhibited linear kinetics up to 90% conversion using 0.5 equivalents of Me6-TREN with respect to initiator concentration.