Poly(l-lysine) grafted with poly(ethylene glycol) (PLL-g-PEG), a polycationic copolymer that is positively charged at neutral pH, spontaneously adsorbs from aqueous solution onto negatively charged surfaces, resulting in the formation of stable polymeric monolayers and rendering the surfaces protein-resistant to a degree related to the PEG surface density. A set of PLL-g-PEG polymers with different architectures was synthesized. The grafting ratio, g, of the polymer, defined as the ratio of the number of lysine monomers to the number of PEG side chains, was systematically varied between 2 and 23, and PEG molecular weights of 1, 2, and 5 kDa were used. The polymers were adsorbed onto niobium oxide-coated substrates, leading to highly different but well-controlled PEG surface densities with maximal values of 0.9, 0.5, and 0.3 chains/nm2 for the three PEG molecular weights, respectively. Time-of-flight secondary-ion mass spectrometry (ToF-SIMS) was used in conjunction with the in situ optical waveguide lightmode spectroscopy (OWLS) technique to investigate the interface architecture. While ToF-SIMS provided surface-analytical data on the polymeric adlayer, OWLS allowed the quantitative determination of the adsorbed polymer mass. Extremely good correlations were established between the ToF-SIMS data (obtained in UHV) and the in situ OWLS results. The amount of serum adsorbed, determined quantitatively by OWLS, was found to depend systematically on the surface coverage in terms of the ethylene glycol (EG) density, controlled by both PEG molecular weight and grafting ratio, g. Serum adsorption dropped gradually from 590 ng/cm2 on bare Nb2O5 to <2 ng/cm2 (=detection limit of the OWLS technique) for EG densities ≥ 20 nm-2. The PLL-g-PEG technology shows itself to be an efficient, cost-effective, and robust tool for the immobilization of PEG chains onto metal oxide surfaces. The precise control over PEG surface density across a wide range allows for the production of tailored surfaces with controlled degrees of bio-interactiveness. Such surfaces are expected to have a substantial potential for applications in biomedical and bioanalytical devices.

A novel biosensor interface exploiting the spontaneous surface assembly of a polycationic, PEG-grafted, biotinylated copolymer was developed and tested on optical waveguide chips in a model immunoassay based on sequential immobilization of (strept)avidin and biotinylated goat antirabbit immunoglobulin (RRIgG-biotin) as a capture molecule to sense the rabbit immunoglobulin (RIgG) target molecule. Optical waveguide lightmode spectroscopy with niobium oxide waveguiding layers was used to monitor quantitatively and in situ the spontaneous adsorption of the (biotinylated) copolymer onto the waveguide surface, the resistance of the resulting adlayer to nonspecific protein adsorption, and the mass uptakes in each step of the model immunoassay. Poly(L-lysine)-g-poly(ethylene oxide) (PLL-g-PEG) is a polycationic copolymer that adsorbs spontaneously from aqueous solutions onto negatively charged surfaces via electrostatic interactions. It forms monolayers with densely packed PEG chains. PLL-g-PEG graft copolymers carrying terminal biotin groups on 0, 20, 30, or 50% of the PEG chains were synthesized and assembled onto the surface of niobium oxide (negatively charged at neutral pH). The surface concentration of biotin was tailored by adjusting the biotin grafting ratio in the polymeric molecule or by assembling mixed [PLLg-PEG/PEGbiotin + PLL-g-PEG] adlayers from the corresponding mixed solutions. These biotinylated surfaces are shown to be highly resistant to nonspecific adsorption from serum while still allowing for the specific surface binding of the linkage proteins: streptavidin, avidin, or neutral avidin. The amount of immobilized linkage protein is shown to be closely related to the biotin surface concentration. The subsequent adsorption behavior of RRIgG-biotin and RIgG, however, depends in a more complex manner on each individual surface modification step and is discussed in the light of specific and nonspecific interactions, as well as of orientational and steric repulsion effects within the adlayers. In terms of the sensing signalto-background ratio, the [PLL-g-PEG/PEGbiotin//NeutrAvidin//RRIgG-biotin] architecture demonstrated particularly promising performance as an interface architecture for bioaffinity sensing of proteins.

No abstract

No abstract