ABSTRACT
A portable and highly sensitive sensor was designed for the specific detection of 3,4-methyl-enedioxy-methamphetamine (MDMA), in a range of field-testing situations. The sensor can detect MDMA in street samples, even when other controlled substances drugs, or adulterants are present. In this work, we report for the first time a sensor using electroactive molecularly imprinted polymer nanoparticles computationally designed to recognize MDMA and then produced using solid phase synthesis. A composite comprising chitosan, reduced graphene oxide, and molecularly imprinted polymer nanoparticles synthesized for MDMA for the first time was immobilized on screen-printed carbon electrodes. The sensors displayed a satisfactory sensitivity (106.8 nA × µM-1 ), limit of detection (1.6 nM; 0.31 ng/mL), and recoveries (92-99%). The accuracy of the results was confirmed through validation using Ultra-High Performance Liquid Chromatography coupled with tandem Mass Spectrometry (UPLC-MS/MS). This technology could be used in forensic analysis and make it possible to selectively detect MDMA in street samples.
ABSTRACT
Cellular senescence has proved to be a strong contributor to ageing and age-related diseases, such as cancer and atherosclerosis. Therefore, the protein content of senescent cells is highly relevant to drug discovery, diagnostics and therapeutic applications. However, current technologies for the analysis of proteins are based on a combination of separation techniques and mass spectrometry, which require handling large sample sizes and a large volume of data and are time-consuming. This limits their application in personalised medicine. An easy, quick and inexpensive procedure is needed for qualitative and quantitative analysis of proteins expressed by a cell or tissue. Here, we describe the use of the "snapshot imprinting" approach for the identification of proteins differentially expressed by senescent cells. Molecularly imprinted polymer nanoparticles (MIPs) were formed in the presence of whole cells. Following trypsinolysis, protein epitopes protected by complex with MIPs were eluted from the nanoparticles and analysed by LC-MS/MS. In this work, "snapshot imprinting" was performed parallel to a standard proteomic "shaving approach", showing similar results. The analysis by "snapshot imprinting" identified three senescent-specific proteins: cell division cycle 7-related protein kinase, partitioning defective three homolog B and putative ATP-dependent RNA helicase DHX57, the abundance of which could potentially make them specific markers of senescence. Identifying biomarkers for the future elimination of senescent cells grants the potential for developing therapeutics for age-related diseases.
ABSTRACT
Molecularly Imprinted Polymers (MIPs) are synthetic receptors capable of selective binding to their target (template) molecules and, hence, are used as recognition elements in assays and sensors as a replacement for relatively unstable enzymes and antibodies. Herein, we describe a manufacturing-friendly protocol for integration of MIP nanoparticles (nanoMIPs) with a (label-free) capacitive sensor. The nanoMIPs were produced by solid-phase synthesis for two templates with different sizes and properties, including a small molecule tetrahydrocannabinol (THC) and a protein (trypsin). NanoMIPs were deposited on the surface of the sensor and the change in capacitance (ΔC) upon binding of the target was measured. The significant improvement in the selectivity and limit of detection (one order of magnitude compared to previously used MIP microparticles) can be attributed to their increased surface-to-volume ratio and higher specificity of the nanoMIPs produced by the solid-phase method. The methodology described is also compatible with common sensor fabrication approaches, as opposed to methods involving in situ MIP polymerisation. The proposed sensor shows high selectivity, fast sensor response (45â¯min including injection, regeneration and re-equilibration with running buffer), and straightforward data analysis, which makes it viable for label-free monitoring in real-time. The set of targets assessed in this manuscript shows the general applicability of the biosensor platform.
Subject(s)
Biosensing Techniques/methods , Nanoparticles/chemistry , Polymers/chemistry , Molecular ImprintingABSTRACT
The enzyme-linked immunosorbent assay (ELISA) has been used as a standard tool for monitoring food and animal feed contamination from the carcinogenic fumonisin B1 (FB1). Unfortunately, ELISA is not always efficient due to the instability of the antibody and enzyme components in the immunoassay, the presence of natural enzyme inhibitors in the samples and the high levels of non-specific protein binding. Additionally, the production of antibodies for ELISA can be time-consuming and costly, due to the involvement of animals in the manufacturing process. To overcome these limiting factors, a molecularly imprinted nanoparticle based assay (MINA) has been developed, where the molecularly imprinted nanoparticles (nanoMIPs) replace the primary antibody used in a competitive ELISA. Herein, computational modelling was used to design the nanoMIPs by selecting monomers that specifically interact with FB1. The affinity of the monomers to FB1 was verified by measuring their binding in affinity chromatography experiments. The nanoMIPs were produced by solid phase synthesis and the results showed that nanoMIPs had a hydrodynamic diameter of around 249 ± 29 nm. The assay tested in model samples is highly selective and does not show cross-reactivity with other mycotoxins such as fumonisin B2 (FB2), aflatoxin B1 (AFB1), citrinin (CTT), zearalenone (ZEA), and deoxynivalenol (DON). The MINA allows the detection of FB1 in the concentration range of 10 pM-10 nM with a detection limit of 1.9 pM and a recovery of 108.13-113.76%.
