Automatic reactor for solid-phase synthesis of molecularly imprinted polymeric nanoparticles (MIP NPs) in water

Abstract

We report the development of an automated chemical reactor for solid-phase synthesis of MIP NPs in water. Operational parameters are under computer control, requiring minimal operator intervention. In this study, "ready for use" MIP NPs with sub-nanomolar affinity are prepared against pepsin A, trypsin and α-amylase in only 4 hours.The extent of research into biomedical and diagnostic nanotechnology is impressive, 1 despite currently very few examples of biomedical nanomaterials have been approved by the FDA. 2 Apart from the potential toxicity, the usage of nanomaterials in therapy and diagnostics finds a strong opponent in the well-established leadership of "safer" natural antibodies. 3 Industrial manufacturing of antibodies, however, is logistically complex and expensive, especially for applications in therapy. 4 Potential immunogenic adverse reactions, low stability and poor performance in non-physiological conditions restrict the application of antibodies to specific niche areas and shorten their shelf-life. 5,6 Finally, natural antibodies can be difficult to integrate with assays and sensors. 6,7 A possible alternative are MIP NPs, which share key characteristics with antibodies such as aqueous solubility, size, affinity and selectivity for the target. Additionally, solutions of MIP NPs can be handled similarly to solutions of antibodies, with the advantage of stability and robustness. 8 Moreover, operational parameters for producing MIP NPs can be carefully controlled. 9 There is, however, lack of a generic protocol for the synthesis of MIP NPs with specific characteristics, 8 which restricts their practical applications, despite recent reports of biological activity 10 and in vivo detoxification 11 hint at a great potential of MIP NPs in the nanomedicine arena.With the aim to provide a method that is scalable, reproducible and controlled, we have focused our efforts on the development of synthetic strategies for producing highperformance MIP NPs. The most suitable protocol should in our view include affinity purification with immobilised template to remove monomers/low-affinity NPs from the product. 12 Even better results would be obtained if the immobilised template is used also in polymerisation, as this will ensure high-binding site affinity and prevent the contamination 13 This approach would also add advantages of easy automation and the potential for scaling-up production.Recently we have reported the first successful example of solid-phase synthesis of MIP NPs using living-polymerisation chemistry, performed in organic solvent under UV irradiationconditions which are favourable for the imprinting of small molecules. 14 Imprinting of highmolecular weight targets such as proteins, polysaccharides and DNA, however, would require an aqueous environment to preserve their structure during polymerisation. 15Here we report the development of a reactor for MIP NPs preparation using chemical polymerisation performed in specifically mild aqueous conditions (monomer concentration: 6.5 mM) for p...

Synthetic Strategies in Molecular Imprinting

“…To solve this problem, one feasible approach is to develop MIPs as modular building blocks that can be handled like antibodies for different applications. In this respect MIP nanoparticles are particularly interesting, as they can be prepared in large quantity with high molecular selectivity [64], and their small physical size allows the material to be easily handled with standard liquid dispensers.…”

Electrosynthesized molecularly imprinted polymers for protein recognition

“…Imprinted soluble nanogels, having dimensions comparable to that of protein clusters were also shown to allow facile template exchange between the polymer and the solution [32]. The so-called solid-phase synthesis approach made feasible the reproducible, controlled and large scale production of such imprinted nanogels [33][34][35]. Interfacial polymerization on planar surfaces or supporting beads [36][37][38] can create ultra-thin polymer films that only partially bury the protein template without completely encapsulating it.…”

Molecular imprinting: perspectives and applications

“…The automatic method offered short synthesis/ purification times, thus making it potentially suitable for industrial applications and production. 134 …”

“…The advantage of the method was that all binding sites had the same orientation and were located at the surface of the glass beads, thus improving binding site homogeneity and accessibility. 133 In 2014, Poma et al 134 developed an automated chemical reactor for solid-phase synthesis of MIP-NPs in water for proteins such as trypsin, pepsin A and a-amylase. The automatic method offered short synthesis/ purification times, thus making it potentially suitable for industrial applications and production.…”