Vitamin D3 (VD3) is an essential micronutrient, but its analytical determination in biological matrices is often hindered by structurally related metabolites and the limited selectivity of conventional analytical sorbents. The preparation of a molecularly imprinted polymer (MIP) using VD3 as a template is challenging due to its hydrophobic structure and lack of polar groups. Therefore, in this work, MIPs were prepared using the closely related structure hyodeoxycholic acid methyl ester as a template and tested for their adsorption capacity toward VD3. Several MIPs were first prepared using different functional monomers, and the results showed that 4-vinylpyridine (4VP) monomer in combination with divinylbenzene (DVB) as a crosslinker exhibited a relatively high binding capacity and imprinting factor. UV spectroscopy indicated an optimal VD3–monomer ratio of 1:4, while computational modeling further confirmed favorable interactions between VD3 and 4VP. The effect of incorporating styrene as a co-monomer with 4VP was also investigated, showing an enhancement in adsorption capacity with a slight increase in the imprinting factor. However, TGA analysis revealed that the thermal stability of the MIPs decreased with higher styrene content. Overall, the prepared MIPs demonstrated improved selectivity and recognition of VD3 compared to the non-imprinted polymers, offering a promising approach for its selective extraction and quantification.

Site-level resolution of protein phosphorylation remains a central challenge in decoding cellular signaling and conventional global phosphoproteomics is limited in its ability to detect low-abundance peptides and resolve positional isomers. Here we introduce sequence-selective synthetic receptors based on imprinted polymers for targeted enrichment of defined phosphopeptide motifs from complex cancer cell proteomes. By encoding local sequence context into the binding interface, these receptors enable selective capture of closely related phosphorylation sites with high specificity. Using the T cell kinase ZAP70 as a model, we resolve phosphorylation at adjacent tyrosine residues (pY492 and pY493) within a regulatory motif, overcoming a long-standing limitation in site discrimination. Integrated with liquid chromatography–mass spectrometry workflows, this enables sensitive detection of low-abundance and isomeric phosphopeptides that evade conventional enrichment strategies. More broadly, our findings establish molecular imprinting as a programmable chemical platform for site-resolved phosphoproteomics, opening new avenues to interrogate signaling networks with molecular precision.

Protein phosphorylation is a reversible post-translational modification (PTM) acting as a key regulator of protein function with an impact on cell behavior and pathological conditions. Accurate monitoring of protein phosphorylation status is important for both mechanistic deciphering of cellular processes and disease diagnostics. However, detecting this PTM remains challenging due to its transient nature, stability issues, and low abundance. Molecularly imprinted polymers (MIPs) fill a void in the search for robust affinity techniques that can be tailored to specifically enrich phospho-peptides at predetermined levels of discrimination. This ranges from (i) surface-imprinted mesoporous materials for generic enrichment; (ii) amino acid–specific receptors that fractionate by side chain with minimal sequence bias; (iii) dummy-templated MIPs for labile phosphorylations; to (iv) sequence-specific receptors for defined phospho-epitopes and biomarkers. This review offers a concise, interdisciplinary overview of the field in the context of current challenges in phosphoproteomics.

The paper reports the design of multivalent bacterial receptors based on reversible self-assembled monolayers (rSAMs) on gold and glass substrates, mimicking the ligand display on host cells and extracellular matrices. The layers consist of α-(4-amidinophenoxy)alkanes decorated at the ω-position with β-galactose (Gal) or sialic acid (SA). The former acts as a mobile ligand binding to the complementary adhesin, LecA, a key virulence factor of the multi-drug-resistant bacterium Pseudomonas aeruginosa (PA). Binary amphiphile mixtures containing either of these ligands, spontaneously self-assemble on carboxylic acid terminated SAMs on gold or glass surfaces to form rSAMs that are easily tunable with respect to the ligand ratio. It is shown that this results in the ability to construct multi-reusable surfaces featuring strong affinity for the bacterial adhesin and recognitive surfaces for bacteria, the latter demonstrated by incubating a culture of PA or the oral commensal species Streptococcus gordonii (SG) on either Gal or SA functionalized rSAMs. In contrast to the mobile ligand display, surfaces featuring covalently attached "static" ligands exhibited low LecA affinity. This approach to wet chemical surface functionalization is unique in imparting both rapid restorability and adaptability, the latter compatible with heteromultivalent receptor designs for boosting lectin and bacteria affinity and specificity.

Diagnostic pregnancy tests are the most widely used immunoassays for home-based use. These tests employ the well-established lateral flow assay (LFA) technique, reminiscent of affinity chromatography relying on the dual action of two orthogonal anti-hCG antibodies. Immunoassays suffer from several drawbacks, including challenges in antibody manufacturing, suboptimal accuracy, and sensitivity to adverse storing conditions. Additionally, LFAs are typically designed for single use, as the LFA technique is non-reusable. An alternative to overcome these drawbacks is to leverage molecularly imprinted polymer (MIP) technology to generate polymer-based hCG-receptors and, subsequently, non-bioreceptor-based tests. Here, we report the development of MIP nanogels for hCG detection, exploiting epitopes and magnetic templates for high-yielding dispersed phase imprinting. The resulting nanogels were designed for orthogonal targeting of two immunogenic epitopes (SV and PQ) and were thoroughly characterized with respect to physical properties, binding affinity, specificity, and sensitivity. Molecular dynamics simulations indicated a pronounced conformational overlap between the templates and the epitopes in the native protein, supporting their suitability for templating cavities for hCG recognition. Quartz crystal microbalance (QCM)-based binding tests and kinetic interaction analysis by surface plasmon resonance (SPR) revealed nanomolar dissociation constants for the MIP nanogels and their corresponding template peptides and low uptake of lutenizing hormone (LH), structurally resembling to hCG. Receptor reusability was demonstrated in the multicycle SPR sensing mode using a low pH regeneration buffer. The results suggest the feasibility of using imprinted nanogels as a class of cost-effective, stable alternatives to natural antibodies for hCG detection. We foresee applications of these binders with respect to reusable pregnancy tests and other hCG-related disease diagnostics.

We report a combinatorial approach to design peptide-based biomimetic sensors for detecting β-type coronaviruses with high sensitivity and selectivity. We selected three peptide epitopes from key regions of the ACE2 receptor that are involved in viral binding to different variants and immobilized them individually or in binary combinations on gold sensor chips. Using Surface Plasmon Resonance (SPR), we found that single-epitope sensors displayed nanomolar dissociation constants to three RBD variants (SARS-CoV-2 Delta < SARS-CoV-2 Alpha < SARS-CoV-1) and a KD = 1.2 ± 0.4 nM to the full SARS-CoV-2 Alpha spike protein, with negligible binding to the a-coronavirus hCoV-NL63 spike protein. Molecular dynamics simulations revealed that the tightest binding epitope closely mimics ACE2 interactions with β-coronaviruses, explaining its superior performance. In contrast, dual epitope systems exhibited a reversed variant preference, with a pronounced affinity enhancement for SARS-CoV-1 (KD = 6 ± 2 nM). This was attributed to cooperative epitope interactions that could restrict the conformational flexibility of the longer epitope, favoring the effective intermolecular contacts that strengthen the interaction with the RBD. These findings suggest a time-saving approach for developing sensitive and selective sensors for rapidly mutating viruses.

Multivalent inhibitors that mimic the polysaccharide array on cells represent a new paradigm in the development of antiviral agents and antibiotics. Covalent ligand anchoring limits the affinity and, in turn, potency of these inhibitors with dissociation constants (K-d) commonly found in the micromolar or upper nanomolar range. Addressing this deficiency we here report on easily accessible gold core-shell nanoparticles (rSAM-NPs) featuring adaptable reversible self-assembled monolayer (rSAM)-based shells. The rSAMs are anchored by noncovalent amidinium-carboxylate interactions on gold nanoparticles at slightly alkaline pH resulting in laterally mobile pH-responsive assemblies that are functional at physiological pH. Introducing sialic acid ligands in the shell, we show that the rSAM-NPs strongly interact with the influenza virus surface protein hemagglutinin (limit of detection LoD < 2 nM) and deactivated bird flu virus H5N1 (LoD < 1 HAU) in allantoic liquid. Finally, we show that the rSAM-NPs effectively inhibit the interaction of the virus with red blood cells at concentrations in the low picomolar range. This represents a significant increase in potency with respect to multivalent inhibitors of similar size based on covalently anchored monosaccharides.

Antibiotic determination at low concentrations is a critical challenge to ensure water, food, and environmental safety. This work reports the application of molecularly imprinted nanoparticles (MINs) as synthetic antibodies in a microplate assay for tetracycline (TC) determination at subnanomolar levels. MINs with hydrodynamic radii ranging from 200 to 300 nm were synthesized via solid-phase imprinting on magnetic nanoparticles using lymecycline (LMC) as an auxiliary template. MINs immobilized onto polystyrene microplates exhibited a selective binding of the conjugate between LMC and horseradish peroxidase (HRP), yielding analytical responses up to fourfold higher than those observed in blank wells. MINs saturated with LMC-HPR showed the quantitative displacement of the conjugate upon incubation with LMC or TC, yielding concentration-proportional responses from 0.001 to 100 nmol L-1. For LMC, we obtained limits of detection (LOD) between 0.018 and 0.020 nmol L-1 and limits of quantification (LOQ) from 0.054 to 0.061 nmol L-1. For TC, the LOD values ranged between 0.001 and 0.006 nmol L-1, with LOQ values of 0.004-0.019 nmol L-1. Within the studied concentration range, the analytical responses of MINs were two to five times higher than those obtained from nonimprinted materials or blank controls. These findings support the performance of MINs as antibody mimetics with high recognition capacity, relatively low production costs, and robust analytical applications.

Prostate-specific membrane antigen (PSMA) is overexpressed in prostate cancer cells and tumor vasculature, making it an important biomarker. However, conventional PSMA-targeting agents like antibodies and small molecules have limitations. Antibodies exhibit instability and complex production, while small molecules show lower specificity and higher toxicity. Herein, this work develops a novel PSMA-targeting synthetic antibody to address prior limitations. This work synthesizes fluorescently labelled, N-isopropylacrylamide-based epitope imprinted nanogels (MIP-M) using a dispersion of magnetic nanoparticles as template carriers with a linear epitope from PSMA's extracellular apical domain as the template. MIP-M demonstrates high binding affinities for both the epitope template (apparent K_D = 6 × 10^-10 м) and PSMA (apparent K_D = 2.5 × 10^-9 м). Compared to reference peptides and human serum albumin, MIP-M indicates high specificity. Flow cytometry and confocal laser scanning microscopy comparing cell lines displaying normal (PC3) and enhanced (LNCaP) PSMA expression levels, revealed that MIP-M and a PSMA antibody exhibits comparable binding preferences for the latter cell line. Moreover, MIP-M demonstrates selectivity on par with the PSMA antibody for targeting PSMA-positive prostate tumor over normal tissue, enabling discrimination. This MIP-M addresses stability, production, specificity and toxicity limitations of prior targeting agents and offer a promising alternative for PSMA-directed cancer diagnosis and treatment. 

Stimuli‐responsive molecularly imprinted polymers (MIPs) are exciting smart materials that are gaining substantial interest within the research community due to their versatility and possible widespread applications in biosensing, biomedicine and diagnostics, as well as chromatography and separation sciences. These materials offer significant advantages as recognition materials over their biological counterparts (antibodies) because of their ease and low cost of production along with their robustness and resistance to the extremes of temperature and pH. This much needed review aims to provide an updated summary of the various stimuli‐responsive MIPs reported to date including those relying on thermo, pH, photo, biomolecule, ion, magnetic and electrical stimuli and includes their design and synthesis. The review also explores the potential applications of the stimuli‐responsive MIPs, particularly in the fields of biosensors and diagnostics, along with biological imaging, drug delivery, disease treatments and interventions and the separation of targets from complex media. The advantages and disadvantages of the current stimuli‐responsive MIPs set out in the review, allows for researchers to gather a concise understanding of these smart‐materials and should pave the way for new methods of development and real‐world applications. We believe the review is a helpful and necessary guide for the future evolution and application of stimuli‐responsive MIPs.