Selection of imprinted nanoparticles by affinity chromatography

Soluble molecularly imprinted nanoparticles were synthesised via iniferter initiated polymerisation and separated by size via gel permeation chromatography. Subsequent fractionation of these particles by affinity chromatography allowed the separation of high affinity fractions from the mixture of nanoparticles. Fractions selected this way possess affinity similar to that of natural antibodies (Kd 6.6 × 10−8) M and were also able to discriminate between related functional analogues of the templ

The rational development of molecularly imprinted polymer-based sensors for protein detection.

The detection of specific proteins as biomarkers of disease, health status, environmental monitoring, food quality, control of fermenters and civil defence purposes means that biosensors for these targets will become increasingly more important. Among the technologies used for building specific recognition properties, molecularly imprinted polymers (MIPs) are attracting much attention. In this critical review we describe many methods used for imprinting recognition for protein targets in polymers and their incorporation with a number of transducer platforms with the aim of identifying the most promising approaches for the preparation of MIP-based protein sensors (277 references)

Substitution of antibodies and receptors with molecularly imprinted polymers in enzyme-linked and fluorescent assays

A new technique for coating microtitre plates with molecularly imprinted polymers (MIP), specific for low-molecular weight analytes (epinephrine, atrazine) and proteins is presented. Oxidative polymerization was performed in the presence of template; monomers: 3-aminophenylboronic acid, 3- thiopheneboronic acid and aniline were polymerized in water and the polymers were grafted onto the polystyrene surface of the microplates. It was found that this process results in the creation of synthetic materials with antibody-like binding properties. It was shown that the MIP-coated microplates are particularly useful for assay development. The high stability of the polymers and good reproducibility of the measurements make MIP coating an attractive alternative to conventional antibodies or receptors used in ELISA

The stabilisation of receptor structure in low cross-linked MIPs by an immobilised template

In molecularly imprinted polymers (MIPs) a high level of cross-linking is usually important for preserving the receptor structure. We propose here an alternative approach for stabilising binding sites, which involves the use of an immobilised template. The idea is based on the assumption that an immobilised template will ‘‘hold’’ polymeric chains and complementary functionalities together, preventing the collapsing of the binding sites. To test this postulate, a range of polymers was prepared using polymerisable (2,4-diamino-6- (methacryloyloxy)ethyl-1,3,5-triazine) and non-polymerisable (or extractable) (2,4-diamino-6-methyl-1,3,5-triazine) templates, methacrylic acid as functional monomer and ethylene glycol dimethacrylate as cross-linker. The level of cross- linking was varied from 12 to 80%. Polymerisations were performed in acetonitrile using UV initiation. Binding properties of the synthesised materials were characterised both by HPLC and equilibrium batch binding experiments followed by HPLC-MS or UV-visible detection. The adsorption isotherms of polymers were obtained and fitted to the Langmuir model to calculate dissociation constant, Kd, and concentration of binding sites for each material. The results strongly indicate that the presence of an immobilised template improves the affinity of MIPs containing low percentages of cross- linker. The low cross-linked MIPs synthesised with a polymerisable template also retain a reasonable degree of selectivity. Low crosslinked MIPs with such binding characteristics would be useful for the creation of new types of optical and electrochemical sensors, where induced fit or the ‘‘gate effect’’ could be used more effectively for generating and enhancin

Utilization of Synthetic Antibody for Fumonisin Determination in Feed and Food

Fumonisin contamination in food is limited around 2 – 4 ppm and in feed for different animals varies from 5 to 100 ppm. This regulation is to prevent animal and human from carcinogenic effect from fumonisins. Measurement of fumonisins frequently uses chromatography methods such as High-Performance Liquid Chromatography (HPLC) and Liquid chromatography tandem-mass spectrometry (LCMS/MS); however, the sample preparation and analysis process for these methods are costly and time consuming. Immunoassays have also been employed for detecting fumonisins in food or feed. Unfortunately, the instability of antibody to harsh condition such as high temperature and pH becomes the drawback for immunoassay method. Currently, the technology based on molecularly imprinting, which is called synthetic antibody, has been established for replacing antibody functions. Therefore, the aim of this review is to describe development of molecularly imprinted polymer (MIP) in fumonisin analysis in feed and food. Herein, the composition and production of MIP were described comprehensively. Bulk polymerization and solid phase synthesis were methods for production of MIP in micro and nano sizes. The application of MIP was reported for sample preparation as solid phase extraction measured continuously by HPLC showing the high recovery (> 60%). Then, MIP replaced antibody in direct competitive enzyme-linked immunosorbent assay (ELISA) for quantifying fumonisins in maize with high recovery (>90%) and limit detection (2 – 6 pM). Lastly, MIP was also employed in electrochemical sensor application as receptor for recognizing fumonisin in milk and maize. In conclusion, the performance of MIP has been applied successfully for fumonisin analysis comprehensively from sample preparation and quantification. The MIP would be developed for wider application for other toxins in feed or food such as veterinary drug, heavy metals, or pesticides

Preliminary evaluation of military, commercial and novel skin decontamination products against a chemical warfare agent simulant (methyl salicylate)

This is an Accepted Manuscript of an article published by Taylor & Francis in Cutaneous and Ocular Toxicology on 13 August 2015, available online: http://www.tandfonline.com/doi/full/10.3109/15569527.2015.1072544.Rapid decontamination is vital to alleviate adverse health effects following dermal exposure to hazardous materials. There is an abundance of materials and products which can be utilised to remove hazardous materials from the skin. In this study, a total of 15 products were evaluated, 10 of which were commercial or military products and 5 were novel (molecular imprinted) polymers. The efficacies of these products were evaluated against a 10µL droplet of 14C-methyl salicylate applied to the surface of porcine skin mounted on static diffusion cells. The current UK military decontaminant (Fuller’s earth) performed well, retaining 83% of the dose over 24 hours and served as a benchmark to compare with the other test products. The five most effective test products were Fuller’s earth (the current UK military decontaminant), Fast-Act® and three novel polymers (based on itaconic acid, 2-trifluoromethylacrylic acid and N,N-methylene bis acrylamide). Five products (medical moist free wipes, 5% FloraFree™ solution, normal baby wipes, baby wipes for sensitive skin and Diphotérine™) enhanced the dermal absorption of 14C-methyl salicylate. Further work is required to establish the performance of the most effective products identified in this study against chemical warfare agents.Peer reviewedFinal Accepted Versio

Catalytic molecularly imprinted polymer membranes: Development of the biomimetic sensor for phenols detection

Portable biomimetic sensor devices for the express control of phenols content in water were developed. The synthetic binding sites mimicking active site of the enzyme tyrosinase were formed in the structure of free-standing molecularly imprinted polymer membranes. Molecularly imprinted polymer membranes with the catalytic activity were obtained by co-polymerization of the complex Cu (II)–catechol–urocanic acid ethyl ester with (tri)ethyleneglycoldimethacrylate, and oligourethaneacrylate. Addition of the elastic component oligourethaneacrylate provided formation of the highly cross-linked polymer with the catalytic activity in a form of thin, flexible, and mechanically stable membrane. High accessibility of the artificial catalytic sites for the interaction with the analyzed phenol molecules was achieved due to addition of linear polymer (polyethyleneglycol Mw 20,000) to the initial monomer mixture before the polymerization. As a result, typical semi-interpenetrating polymer networks (semi-IPNs) were formed. The cross-linked component of the semi-IPN was represented by the highly cross-linked catalytic molecularly imprinted polymer, while the linear one was represented by polyethyleneglycol Mw 20,000. Extraction of the linear polymer from the fully formed semi-IPN resulted in formation of large pores in the membranes’ structure. Concentration of phenols in the analyzed samples was detected using universal portable device oxymeter with the oxygen electrode in a close contact with the catalytic molecularly imprinted polymer membrane as a transducer. The detection limit of phenols detection using the developed sensor system based on polymers–biomimics with the optimized composition comprised 0.063 mM, while the linear range of the sensor comprised 0.063–1 mM. The working characteristics of the portable sensor devices were investigated. Storage stability of sensor systems at room temperature comprised 12 months (87%). As compared to traditional methods of phenols detection the developed sensor system is characterized by simplicity of operation, compactness, an

Development of a molecularly imprinted polymer specific for ochratoxin A : theoretical and sensor applications

In this work the development of two molecularly imprinted polymers, specific for ochratoxin A, is presented. Ochratoxin A is produced by several Aspergillus and Penicillium species and is common in cereals and other starch rich foods and has also been found in coffee, dried fruits, wine, beer and meats. It demonstrates potent teratogenic, immunosuppressive, mutagenic and carcinogenic properties. The toxin is also linked to Balkan Endemic Nephropathy, a chronic kidney disease found in South-Eastern Europe. Due to this the European Union has set limits on foodstuffs ranging between 2-10 ng g-1. Therefore the requirement of a simple and inexpensive biosensor to monitor this legislation is a necessity. Currently detection is performed by chromatographic methods such as HPLC, and by ELISA formats. In this work two polymeric materials, rationally designed by computational modelling and synthesised using molecular imprinting, are studied. The modelling is complimented with a Nuclear Magnetic Resonance (NMR) study. The first polymer (Polymer A) consisted of 1 mol of acrylamide and 1 mol of methacrylic acid to 1 mol of template. This material demonstrated an unusual binding mechanism, working solely in aqueous solvents. A theoretical mechanism for this binding is presented and discussed. The second polymer, again rationally designed, but under different conditions, consisted of 1 mol of N,N- diethylamino ethyl methacrylate (DEAEM) to 1 mol of template. This polymer demonstrated high affinity for the template in acetonitrile. Polymer A is used in combination with an ion-exchange SPE protocol (developed for this purpose) for the extraction of OTA from maize. Both polymer compositions are used in development of a MIP membrane optical sensor, with partial success seen in the detection of OTA in grape juice and white wine.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Biomimetic sensors for HbA1c

Diabetes mellitus is a growing health problem worldwide. Suitable long-term control and management of this disease are enabled by determination of glycated haemoglobin (HbA1c) in blood. The results are given as %HbA1c of total haemoglobin. Presently available tests vary in cost and convenience and there is an identified need to introduce improved equipment for self-monitoring. This dissertation focuses on fast and straightforward detection of glycated haemoglobin (HbA1c) using cyclic voltammetry and chronoamperometry. Haemoglobin was determined by monitoring its reaction with potassium ferricyanide on screen printed electrodes at an oxidative potential +500 mV. A working electrode was modified with carbon nanotubes to enhance electron transfer. A calibration curve was linear in a range from 0.83 to 83 mg/mL. Another innovative approach to detecting haemoglobin using its enzymatic activity was also developed. Detection of haemoglobin was performed with hydroquinone and hydrogen peroxide on screen printed electrodes at a potential -400 mV in a Flow Injection Analysis system (FIA). Cont/d.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Direct replacement of antibodies with molecularly imprinted polymer (MIP) nanoparticles in ELISA - development of a novel assay for vancomycin

A simple and straightforward technique for coating microplate wells with molecularly imprinted polymer nanoparticles (nanoMIPs) to develop ELISA type assays is presented here for the first time. NanoMIPs were synthesized by a solid phase approach with immobilized vancomycin (template) and characterized using Biacore 3000, dynamic light scattering and electron microscopy. Immobilization, blocking and washing conditions were optimized in microplate format. The detection of vancomycin was achieved in competitive binding experiments with a HRP-vancomycin conjugate. The assay was capable of measuring vancomycin in buffer and in blood plasma within the range 0.001-70 nM with a detection limit of 0.0025 nM (2.5 pM). The sensitivity of the assay was three orders of magnitude better than a previously described ELISA based on antibodies. In these experiments nanoMIPs have shown high affinity and minimal interference from blood plasma components. Immobilized nanoMIPs were stored for 1 month at room temperature without any detrimental effects to their binding properties. The high affinity of nanoMIPs and the lack of a requirement for cold chain logistics make them an attractive alternative to traditional antibodies used in ELIS