Metallic nano carrier complex targeting neuroendocrine prostate cancer

Poster presented at Cranfield University’s 2019 Manufacturing Doctoral Community event

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

Molecularly imprinted nanoparticles based sensor for cocaine detection

The development of a sensor based on molecularly imprinted polymer nanoparticles (nanoMIPs) and electrochemical impedance spectroscopy (EIS) for the detection of trace levels of cocaine is described in this paper. NanoMIPs for cocaine detection, synthesized using a solid phase, were applied as the sensing element. The nanoMIPs were first characterized by Transmission Electron Microscopy (TEM) and Dynamic Light Scattering and found to be ~148.35 ± 24.69 nm in size, using TEM. The nanoMIPs were then covalently attached to gold screen-printed electrodes and a cocaine direct binding assay was developed and optimized, using EIS as the sensing principle. EIS was recorded at a potential of 0.12 V over the frequency range from 0.1 Hz to 50 kHz, with a modulation voltage of 10 mV. The nanoMIPs sensor was able to detect cocaine in a linear range between 100 pg mL−1 and 50 ng mL−1 (R2 = 0.984; p-value = 0.00001) and with a limit of detection of 0.24 ng mL−1 (0.70 nM). The sensor showed no cross-reactivity toward morphine and a negligible response toward levamisole after optimizing the sensor surface blocking and assay conditions. The developed sensor has the potential to offer a highly sensitive, portable and cost-effective method for cocaine detection

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)

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

Development of electrochemical immunosensors for HER-1 and HER-2 analysis in serum for breast cancer patients

In this work, two human epidermal growth factor receptors, HER-1 and HER-2, were selected as biomarkers to enable the detection of breast cancer. Therefore, two biosensors were developed using gold sensor chips coupled with amperometric detection of the enzyme label horse radish peroxidase (HRP). The biosensors/immunosensors relied on indirect sandwich enzyme-linked immunosorbent assays with monoclonal antibodies (Ab) against HER-1 and HER-2 attached to the sensors to capture the biomarkers. Detection polyclonal antibodies followed by secondary anti-rabbit (for HER-1) and anti-goat (for HER-2) IgG antibody-HRP were then applied for signal generation. In buffer, the developed sensors showed limits of detections (LOD) of 1.06 ng mL−1 and 0.95 ng mL−1 and limits of quantification (LOQ) of 2.1 ng mL−1 and 1.5 ng mL−1 for HER-1 and HER-2, respectively. In 100% (undiluted) serum, LODs of 1.2 ng mL−1 and 1.47 ng mL−1 and LOQs of 1.5 ng mL−1 and 2.1 ng mL−1 were obtained for HER-1 and HER-2, respectively. Such limits of detections are within the serum clinical range for the two biomarkers. Furthermore, gold nanoparticles (AuNP) labelled with secondary anti-rabbit and anti-goat IgG antibody-HRP were then used to enhance the assay signal and increase the sensitivity. In buffers, LODs of 30 pg mL−1 were seen for both sensors and LOQs of 98 pg mL−1 and 35 pg mL−1 were recorded for HER-1 and HER-2, respectively. For HER-2 the AuNPs biosensor was also tested in 100% serum obtaining a LOD of 50 pg mL−1 and a LOQ of 80 pg mL−1. The HER-2 AuNP electrochemical immunosensor showed high specificity with very low cross-reactivity to HER-1. These findings demonstrate that the two developed sensors can enable early detection as well as monitoring of disease progression with a beneficial impact on patient survival and clinical outcomes

Surface engineered iron oxide nanoparticles generated by inert gas condensation for biomedical applications

Despite the lifesaving medical discoveries of the last century, there is still an urgent need to improve the curative rate and reduce mortality in many fatal diseases such as cancer. One of the main requirements is to find new ways to deliver therapeutics/drugs more efficiently and only to affected tissues/organs. An exciting new technology is nanomaterials which are being widely investigated as potential nanocarriers to achieve localized drug delivery that would improve therapy and reduce adverse drug side effects. Among all the nanocarriers, iron oxide nanoparticles (IONPs) are one of the most promising as, thanks to their paramagnetic/superparamagnetic properties, they can be easily modified with chemical and biological functions and can be visualized inside the body by magnetic resonance imaging (MRI), while delivering the targeted therapy. Therefore, iron oxide nanoparticles were produced here with a novel method and their properties for potential applications in both diagnostics and therapeutics were investigated. The novel method involves production of free standing IONPs by inert gas condensation via the Mantis NanoGen Trio physical vapor deposition system. The IONPs were first sputtered and deposited on plasma cleaned, polyethylene glycol (PEG) coated silicon wafers. Surface modification of the cleaned wafer with PEG enabled deposition of free-standing IONPs, as once produced, the soft-landed IONPs were suspended by dissolution of the PEG layer in water. Transmission electron microscopic (TEM) characterization revealed free standing, iron oxide nanoparticles with size < 20 nm within a polymer matrix. The nanoparticles were analyzed also by Atomic Force Microscope (AFM), Dynamic Light Scattering (DLS) and NanoSight Nanoparticle Tacking Analysis (NTA). Therefore, our work confirms that inert gas condensation by the Mantis NanoGen Trio physical vapor deposition sputtering at room temperature can be successfully used as a scalable, reproducible process to prepare free-standing IONPs. The PEG- IONPs produced in this work do not require further purification and thanks to their tunable narrow size distribution have potential to be a powerful tool for biomedical applications

Development of functionalized nanostructured polymeric membranes for water purification

Pharmaceuticals specific molecularly imprinted polymers nanoparticles (MIPNPs) were synthesized and applied onto the polyvinylidene fluoride (PVDF) membranes previously subjected to the plasma treatment. Diclofenac-, metoprolol- and vancomycin-MIPs were applied onto the membranes and scanning electron microscopy was employed to visualize MIPNPs on the membrane. After functionalization of the membranes with target-specific MIPs the molecularly imprinted membranes (MIMs) affinity against their targets was evaluated using solid phase extraction (SPE) technique coupled with high performance liquid chromatography (HPLC). MIMs were used as filters to load the target solutions through employing a vacuum pump to evaluate the amount of pharmaceuticals in filtrate. Moreover, a comparative study was performed by comparing the efficiency of MIMs functionalized either by adsorption or covalent immobilization. The capacity analysis of MIPNPs by SPE–HPLC revealed 100%, 96.3%, and 50.1% uptake of loaded solution of metoprolol, diclofenac and vancomycin, respectively. MIMs showed 99.6% uptake with a capacity of 60.39 ng cm2 for metoprolol; 94.7% uptake with a capacity of 45.09 ng cm2 for diclofenac; and 42.6% uptake with a capacity of 16.9 ng cm2 for vancomycin. HPLC detection limits of targets were found as 3.7, 7.5 and 15 ng mL−1 for diclofenac, metoprolol and vancomycin respectively. A small scale pilot test was also conducted which indicates the promising future applications of the developed MIMs for high volume of filtrates especially in the case of the plasma-treated PVDF membranes prepared by covalent immobilization of the MIPs

A low-cost miniature immunosensor for haemoglobin as a device for the future detection of gastrointestinal bleeding

Gastrointestinal bleeding (GIB) is a serious medical condition, which requires immediate attention to establish the cause of the bleeding. Here, we present the development of a miniaturised electrochemical impedance spectroscopy (EIS) device for the detection of GIB. The device performs EIS measurements up to 100 kHz. Following the development of an immunosensor for haemoglobin (Hb) on screen printed electrodes, the EIS device was used for detecting Hb as an early indication of bleeding. The sensor was able to detect Hb in a redox solution in a linear range between 5 μg mL−1 and 60 μg mL−1, with a limit of detection of 13.3 μg mL−1. It was also possible to detect Hb in simulated intestinal fluid, without the need for a redox solution, within a range of 10 μg mL−1 to 10 mg mL−1 with a limit of detection of 2.31 mg mL−1. The miniature EIS device developed in this work is inexpensive, with an estimated cost per unit of £30, and has shown a comparable performance to existing commercial tools, demonstrating its potential to be used in the future as an ingestible sensor to detect GIB. All these measurements were carried out in a purpose built flow cell with supporting hardware electronics outside the cell. Integration of the hardware and the sensing electrodes was demonstrated in pill form. This pill after integration sampling fluidics has potential to be used in detecting gastrointestinal bleeding

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