New approach in multipurpose optical diagnostics : fluorescence based assay for simultaneous determination of physicochemical parameters

The development of sensors assays for comprehensive characterisation of biological samples and effective minimal-invasive diagnostics is highly prioritised. Last decade this research area has been actively developing due to possibility of simultaneous, real- time, in vivo detection and monitoring of diverse physicochemical parameters and analytes. The new approach which has been introduced in this thesis was to develop and examine an optical diagnostic assay consisting of a mixture of environmental-sensitive fluorescent dyes. The operating principle of the system has been inspired by electronic nose and tongue devices which combine nonspecific (or semispecific) sensing elements and chemometric techniques for multivariate data analysis. The performance of the optical assay was based on the analysis of the spectrum of selected dyes with discreet reading of their emission maxima. The variations in peaks intensities caused by environmental changes provided distinctive fluorescence patterns, which could be handled similar to the signals collected from nose/tongue devices. The analytical capability of the assay was engendered by changes in fluorescence signal of the dye mixture in response to changes in pH, temperature, ionic strength and the presence of oxygen. Further findings have also proved the ability of optical assay to estimate development phases and to discriminate between different strains of growing cell cultures as well as identify various gastrointestinal diseases in human. This novel fluorescence-based diagnostic tool offers a promising alternative to electrochemical systems providing high sensitive measurements with broad dynamic range, easy, inexpensive measurements and the possibility of remote sensing and extreme assay miniaturisation. Additionally it does not require reference signal. This new approach can impact on a number of applications such as routine minimal- invasive diagnostics for medical samples, biomedical analysis, pharmaceutical or cosmetic research, quality control and process monitoring of food or environmental samples.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

Introducing MINA-The Molecularly Imprinted Nanoparticles Assay

A new ELISA‐ (enzyme‐linked immunosorbent assay)‐like assay is demonstrated in which no elements of biological origin are used for molecular recognition or signaling. Composite imprinted nanoparticles that contain a catalytic core and which are synthesized by using a solid‐phase approach can simultaneously act as recognition/signaling elements, and be used with minimal modifications to standard assay protocols. This assay provides a new route towards replacement of unstable biomolecules in immunoassays

Does size matter? Study of performance of pseudo-ELISAs based on molecularly imprinted polymer nanoparticles prepared for analytes of different sizes

The aim of this work is to evaluate whether the size of the analyte used as template for the synthesis of molecularly imprinted polymer nanoparticles (nanoMIPs) can affect their performance in pseudo-enzyme linked immunosorbent assays (pseudo-ELISAs). Successful demonstration of a nanoMIPs-based pseudo-ELISA for vancomycin (1449.3 g mol) was demonstrated earlier. In the present investigation, the following analytes were selected: horseradish peroxidase (HRP, 44 kDa), cytochrome C (Cyt C, 12 kDa) biotin (244.31 g mol) and melamine (126.12 g mol). NanoMIPs with a similar composition for all analytes were synthesised by persulfate-initiated polymerisation in water. In addition, core-shell nanoMIPs coated with polyethylene glycol (PEG) and imprinted for melamine were produced in organics and tested. The polymerisation of the nanoparticles was done using a solid-phase approach with the correspondent template immobilised on glass beads. The performance of the nanoMIPs used as replacement for antibodies in direct pseudo-ELISA (for the enzymes) and competitive pseudo-ELISA for the smaller analytes was investigated. For the competitive mode we rely on competition for the binding to the nanoparticles between free analyte and corresponding analyte-HRP conjugate. The results revealed that the best performances were obtained for nanoMIPs synthesised in aqueous media for the larger analytes. In addition, this approach was successful for biotin but completely failed for the smallest template melamine. This problem was solved using nanoMIP prepared by UV polymerisation in an organic media with a PEG shell. This study demonstrates that the preparation of nanoMIP by solid-phase approach can produce material with high affinity and potential to replace antibodies in ELISA tests for both large and small analytes. This makes this technology versatile and applicable to practically any target analyte and diagnostic field

Fetal growth trajectory in type 1 pregestational diabetes (PGDM) — an ultrasound study

Objectives: Growth disorders are frequent in diabetic pregnancies. However, they are difficult to predict and capture earlyduring pregnancy. These newborns are at risk of obesity, diabetes, and cardiovascular disease. While developing, fetalgrowth abnormalities are typically progressive. Therefore, capturing the earliest moment when they emerge is essentialto guide subsequent obstetric management.Material and methods: We aimed to analyze fetal ultrasound growth trajectories in type 1 diabetics. Moreover, we aimedto establish time points when first ultrasound manifestations of fetal growth abnormalities appear and to identify factorsthat affect fetal growth in women with diabetes.We collected clinical and ultrasound data from 200 patients with PGDM managed in the third-referential centre for diabetesin pregnancy. During every visit, patients underwent an ultrasound examination according to a standard protocol giving1072 ultrasound scan’s records. Every ultrasound consisted of fetal weight estimation, according to the Hadlock 3 formula.Retrospectively patients were divided into three groups depending on neonatal weight. In the group of 200 patients,60 (30%) delivered LGA and 9 (4.5%) SGA newborns.Results: Fetal growth trajectories show different patterns among fetuses with growth abnormalities in women withtype 1 diabetes. The moment, when fetal growth curves diverge, seems to take place in the second trimester, just afterthe 23rd week of gestation.Conclusions: It suggests that fetal growth abnormalities in type 1 diabetes may have its roots much earlier than expected.In the first trimester, there were differences in LDL-cholesterol, total cholesterol, triglyceride levels and in insulin requirementsbetween AGA, SGA and LGA subgroups

New approach in multipurpose optical diagnostics: fluorescence based assay for simultaneous determination of physicochemical parameters

The development of sensors assays for comprehensive characterisation of biological samples and effective minimal-invasive diagnostics is highly prioritised. Last decade this research area has been actively developing due to possibility of simultaneous, real- time, in vivo detection and monitoring of diverse physicochemical parameters and analytes. The new approach which has been introduced in this thesis was to develop and examine an optical diagnostic assay consisting of a mixture of environmental-sensitive fluorescent dyes. The operating principle of the system has been inspired by electronic nose and tongue devices which combine nonspecific (or semispecific) sensing elements and chemometric techniques for multivariate data analysis. The performance of the optical assay was based on the analysis of the spectrum of selected dyes with discreet reading of their emission maxima. The variations in peaks intensities caused by environmental changes provided distinctive fluorescence patterns, which could be handled similar to the signals collected from nose/tongue devices. The analytical capability of the assay was engendered by changes in fluorescence signal of the dye mixture in response to changes in pH, temperature, ionic strength and the presence of oxygen. Further findings have also proved the ability of optical assay to estimate development phases and to discriminate between different strains of growing cell cultures as well as identify various gastrointestinal diseases in human. This novel fluorescence-based diagnostic tool offers a promising alternative to electrochemical systems providing high sensitive measurements with broad dynamic range, easy, inexpensive measurements and the possibility of remote sensing and extreme assay miniaturisation. Additionally it does not require reference signal. This new approach can impact on a number of applications such as routine minimal- invasive diagnostics for medical samples, biomedical analysis, pharmaceutical or cosmetic research, quality control and process monitoring of food or environmental samples

Optical assay for biotechnology and clinical diagnosis

In this paper, we present an optical diagnostic assay consisting of a mixture of environmental-sensitive fluorescent dyes combined with multivariate data analysis for quantitative and qualitative examination of biological and clinical samples. The performance of the assay is based on the analysis of spectrum of the selected fluorescent dyes with the operational principle similar to electronic nose and electronic tongue systems. This approach has been successfully applied for monitoring of growing cell cultures and identification of gastrointestinal diseases in humans

The Major Hypotheses of Alzheimer’s Disease: Related Nanotechnology-Based Approaches for Its Diagnosis and Treatment

Alzheimer’s disease (AD) is a well-known chronic neurodegenerative disorder that leads to the progressive death of brain cells, resulting in memory loss and the loss of other critical body functions. In March 2019, one of the major pharmaceutical companies and its partners announced that currently, there is no drug to cure AD, and all clinical trials of the new ones have been cancelled, leaving many people without hope. However, despite the clear message and startling reality, the research continued. Finally, in the last two years, the Food and Drug Administration (FDA) approved the first-ever medications to treat Alzheimer’s, aducanumab and lecanemab. Despite researchers’ support of this decision, there are serious concerns about their effectiveness and safety. The validation of aducanumab by the Centers for Medicare and Medicaid Services is still pending, and lecanemab was authorized without considering data from the phase III trials. Furthermore, numerous reports suggest that patients have died when undergoing extended treatment. While there is evidence that aducanumab and lecanemab may provide some relief to those suffering from AD, their impact remains a topic of ongoing research and debate within the medical community. The fact is that even though there are considerable efforts regarding pharmacological treatment, no definitive cure for AD has been found yet. Nevertheless, it is strongly believed that modern nanotechnology holds promising solutions and effective clinical strategies for the development of diagnostic tools and treatments for AD. This review summarizes the major hallmarks of AD, its etiological mechanisms, and challenges. It explores existing diagnostic and therapeutic methods and the potential of nanotechnology-based approaches for recognizing and monitoring patients at risk of irreversible neuronal degeneration. Overall, it provides a broad overview for those interested in the evolving areas of clinical neuroscience, AD, and related nanotechnology. With further research and development, nanotechnology-based approaches may offer new solutions and hope for millions of people affected by this devastating disease

Impact of selected risk factors on motor performance in the third month of life and motor development in the ninth month

Background Proper motor development can be influenced by a range of risk factors. The resulting motor performance can be assessed through quantitative and qualitative analysis of posture and movement patterns. Methods This study was designed as the cohort follow-up of the motor assessment and aimed to demonstrate, in a mathematical way, the impact of particular risk factors on elements of motor performance in the 3rd month and the final motor performance in the 9th month of life. Four hundred nineteen children were assessed (236 male and 183 female), including 129 born preterm. Each child aged 3 month underwent a physiotherapeutic assessment of the quantitative and qualitative development, in the prone and supine positions. The neurologist examined each child aged 9 month, referring to the Denver Development Screening Test II and assessing reflexes, muscle tone and symmetry. The following risk factors were analyzed after the neurological consultation: condition at birth (5th min Apgar score), week of gestation at birth, intraventricular hemorrhage, respiratory distress syndrome, and the incidence of intrauterine hypotrophy and hyperbilirubinemia determined based on medical records. Results A combination of several risk factors affected motor development stronger than any one of them solely, with Apgar score, hyperbilirubinemia, and intraventricular hemorrhage exhibiting the most significant impact. Conclusions Premature birth on its own did not cause a substantial delay in motor development. Nonetheless, its co-occurrence with other risk factors, namely intraventricular hemorrhage, respiratory distress syndrome, and hyperbilirubinemia, notably worsened motor development prognosis. Moreover, improper position of the vertebral column, scapulae, shoulders, and pelvis in the third month of life may predict disturbances in further motor development