A novel method for preparation of biomacromolecular imprinted nanoparticles is described. Combinations of functional monomers were polymerized in the presence of the imprinting peptide melittin in aqueous solution at room temperature to produce a small library of polymer nanoparticles. The template peptide and unreacted monomers are subsequently removed by dialysis. Nanoparticles (NPs) from the library were evaluated for their binding to melittin by 27 MHz QCM analysis. NPs prepared with optimized functional monomer combinations bind strongly to the target molecule. Nanoparticles that were polymerized in the absence of template peptide were found to have little affinity to the peptide. Binding affinity and the size of imprinted particles are comparable to those of natural antibodies. They interact specifically with the target peptide and show little affinity for other proteins. These NPs are of interest as inert and stable substitutes for antibodies. Extension of this approach to other targets of biological importance and the applications of these materials are currently being evaluated.

We report that simple, synthetic organic polymer nanoparticles (NPs) can capture and clear a target peptide toxin in the bloodstream of living mice. The protein-size polymer nanoparticles, with a binding affinity and selectivity comparable to natural antibodies, were prepared by combining a functional monomer optimization strategy with molecular imprinting nanoparticle synthesis. As a result of binding and removal of melittin by NPs in vivo, mortality and peripheral toxic symptoms of melittin were significantly diminished. In vivo imaging of the polymer nanoparticles or "plastic antibodies" establishes the NPs accelerate clearance of the peptide from blood where they accumulate in the liver. Coupled with their biocompatibility and nontoxic characteristics, plastic antibodies offer potential for neutralizing a wide range of biomacromolecules in vivo.In nature, antibodies recognize target molecules by a combination of multiple weak electrostatic, hydrophobic and hydrogen bonding interactions between complementary threedimensional surfaces. To mimic these interactions, nanoparticles (NPs) with affinity for a target peptide or protein have been synthesized by optimizing the composition and ratio of functional groups that make up the NPs.1 , 2 However, the specificity and affinity of the random yhoshino@uci.edu; kjshea@uci.edu. Supporting Information Available: Experimental procedures and supporting data. This material is available free of charge via the Internet at http://pubs.acs.org. We have developed methods for synthesizing protein-size polymer particles with a binding affinity and selectivity comparable to natural antibodies by combining molecular imprinting nanoparticle synthesis with a functional monomer optimization strategy (Figure 1).9 The first stage of this process involves screening small libraries of NPs that span a compositional space chosen for its complementarity to the biological target. 2 The affinity of each NP to the biological target is evaluated and the composition of subsequent NP generations is adjusted to enhance specificity. At the final stage the optimized combination and ratio of functional monomers are polymerized in the presence of the imprinting biological target (peptide or epitope). 9 Following extensive dialysis, polymer NPs exhibit binding affinity, selectivity and particle size comparable to natural antibodies in vitro. NIH Public AccessAlthough molecular recognition by imprinted materials has been extensively studied in controlled settings, little is reported about their application in the bloodstream of living animals. 10 It is well known that the performance (affinity, specificity and function) of synthetic materials when introduced into a complex biological milieu can be profoundly compromised. Introduction of foreign substances including synthetic NPs into the bloodstream results in the immediate formation of a "corona" of proteins on the surface that can alter and/or suppress the intended function of the NP. 11 Further complications can arise fron an immunogenic re...

Synthetic polymer nanoparticles (NPs) that bind venomous molecules and neutralize their function in vivo are of significant interest as "plastic antidotes." Recently, procedures to synthesize polymer NPs with affinity for target peptides have been reported. However, the performance of synthetic materials in vivo is a far greater challenge. Particle size, surface charge, and hydrophobicity affect not only the binding affinity and capacity to the target toxin but also the toxicity of NPs and the creation of a "corona" of proteins around NPs that can alter and or suppress the intended performance. Here, we report the design rationale of a plastic antidote for in vivo applications. Optimizing the choice and ratio of functional monomers incorporated in the NP maximized the binding affinity and capacity toward a target peptide. Biocompatibility tests of the NPs in vitro and in vivo revealed the importance of tuning surface charge and hydrophobicity to minimize NP toxicity and prevent aggregation induced by nonspecific interactions with plasma proteins. The toxin neutralization capacity of NPs in vivo showed a strong correlation with binding affinity and capacity in vitro. Furthermore, in vivo imaging experiments established the NPs accelerate clearance of the toxic peptide and eventually accumulate in macrophages in the liver. These results provide a platform to design plastic antidotes and reveal the potential and possible limitations of using synthetic polymer nanoparticles as plastic antidotes.

Designed polymer nanoparticles (NPs) capable of binding and neutralizing a biomacromolecular toxin are prepared. A library of copolymer NPs is synthesized from combinations of functional monomers. The binding capacity and affinity of the NPs are individually analyzed. NPs with optimized composition are capable of neutralizing the toxin even in a complex biological milieu. It is anticipated that this strategy will be a starting point for the design of synthetic alternatives to antibodies.