Ultrasensitive ELISA using enzyme-loaded nanospherical brushes as labels.

Improving the detection sensitivity of enzyme-linked immunosorbent assay (ELISA) is of utmost importance for meeting the demand of early disease diagnosis. Herein we report an ultrasensitive ELISA system using horseradish peroxidase (HRP)-loaded nanospherical poly(acrylic acid) brushes (SPAABs) as labels. HRP was covalently immobilized in SPAABs with high capacity and activity via an efficient "chemical conjugation after electrostatic entrapment" (CCEE) process, thus endowing SPAABs with high amplification capability as labels. The periphery of SPAAB-HRP was further utilized to bind a layer of antibody with high density for efficient capture of analytes owing to the three-dimensional architecture of SPAABs. Using human chorionic gonadotrophin (hCG) as a model analyte, the SPAAB-amplified system drastically boosted the detection limit of ELISA to 0.012 mIU mL(-1), a 267-fold improvement as compared to conventional ELISA systems.

Covalent immobilization of proteins on 3D poly(acrylic acid) brushes: mechanism study and a more effective and controllable process.

Polymeric brushes have emerged as a novel 3D material platform that provides great amounts of binding sites for biomolecules. This paper investigates the covalent immobilization mechanism of protein by spherical poly(acrylic acid) brushes (SPAABs) in the widely adopted N-hydroxysuccinimide/N-(3-dimethyl-aminopropyl)-N'-ethylcarbodiimide hydrochloride (NHS/EDC) process. It was discovered that electrostatic interaction plays a crucial role in the covalent immobilization of protein. Due to the existence of 3D architecture and "Donnan effect", SPAABs exhibit quite different immobilization kinetics in comparison with conventional 2D materials. Under conditions favorable to electrostatic interaction, the effect of "electrostatic interaction induced covalent binding" was observed as a result of competitive immobilization by physical adsorption and chemical binding. On the basis of the mechanism study, a new "chemical conjugation after electrostatic entrapment" (CCEE) method was developed which set the chemical and physical immobilization process apart. A more effective and well-defined covalent immobilization was achieved. And the binding capacity can be tuned in a wide range (0-4.2 mg protein/mg SPAABs) with a high level of control.

Synthesis and Biomedical Applications of Poly((meth)acrylic acid) Brushes.

Poly((meth)acrylic acid) (P(M)AA) brushes possess a number of distinctive properties that are particularly attractive for biomedical applications. This minireview summarizes recent advances in the synthesis and biomedical applications of P(M)AA brushes and brushes containing P(M)AA segments. First, we review different surface-initiated polymerization (SIP) methods, with a focus on recent progress in the surface-initiated controlled/living radical polymerization (SI-CLRP) techniques used to generate P(M)AA brushes with a tailored structure. Next, we discuss biomolecule immobilization methods for P(M)AA brushes, including physical adsorption, covalent binding, and affinity interactions. Finally, typical biomedical applications of P(M)AA brushes are reviewed, and their performance is discussed based on their unique properties. We conclude that P(M)AA brushes are promising biomaterials, and more potential biomedical applications are expected to emerge with the further development of synthetic techniques and increased understanding of their interactions with biological systems.

A facile route to the synthesis of spherical poly(acrylic acid) brushes via RAFT polymerization for high-capacity protein immobilization.

Spherical poly(acrylic acid) brushes were prepared via a facile RAFT polymerization route from silica nanoparticles (SiO2@PAAs). A silane functionalized RAFT chain transfer agent was designed and synthesized by a one-step reaction, and then immobilized onto silica nanoparticles (SiNPs) through its R group to afford RAFT polymerization. Key structural parameters and contents of carboxyl groups of SiO2@PAAs were thoroughly characterized by transmission electron microscopy, dynamic light scattering, gel permeation chromatography, thermogravimetric analysis and conductometric titration. The SiO2@PAAs exhibit excellent dispersity, tunable brush thicknesses (14.6-68.8 nm) and abundant carboxyl groups (0.82-2.37 mmol/g). An ultra-high protein immobilization capacity (2600 µg streptavidin/mg SiO2@PAAs) was realized by virtue of its rich carboxyl groups and spherical brush structure, which opens up new possibilities for biomedical applications.