A Rapid and Ultra-sensitive Biosensing Platform based on Tunable Dielectrophoresis for Robust POC Applications

With the ongoing pandemic, there have been increasing concerns recently regarding major public health issues such as abuse of organophosphorus compounds, pathogenic bacterial infections, and biosecurity in agricultural production. Biosensors have long been considered a kernel technology for next-generation diagnostic solutions to improve food safety and public health. Significant amounts of effort have been devoted to inventing novel sensing mechanisms, modifying their designs, improving their performance, and extending their application scopes. However, the reliability and selectivity of most biosensors still have much to be desired, which holds back the development and commercialization of biosensors, especially for on-site and point-of-care (POC) usages. Herein, we introduce an innovative two-phase sensing strategy based on tunable AC electrokinetics and capacitive sensing. By dividing the detection process into a sensitivity-priority step and a selectivity-priority step, the specificity and sensitivity of a biosensor can be significantly improved. A capacitive POC aptasensor is fabricated for the implementation of the 2-phase detection and a quasi-single-cell level detection of limit together with an excellent selectivity is achieved simultaneously. The sensor is capable of handling real-world clinic samples without sophisticated pretreatment. Just after a simple one-step dilution, the developed sensor can detect bacteria no less than 2~3 bacteria/10 µL in raw milk samples within 100 s. Alongside whole bacteria detection, the biosensor can also detect endotoxin, the lipopolysaccharide, in bovine serum samples, with a limit of detection of 10 pg/mL. The biosensor is low-cost and easy to use. This work not only demonstrates a biosensor with significant advantages in sensitivity, selectivity and assay time but also opens up a new horizon for further research of all affinity-based biosensors

Nanomaterials for Healthcare Biosensing Applications

In recent years, an increasing number of nanomaterials have been explored for their applications in biomedical diagnostics, making their applications in healthcare biosensing a rapidly evolving field. Nanomaterials introduce versatility to the sensing platforms and may even allow mobility between different detection mechanisms. The prospect of a combination of different nanomaterials allows an exploitation of their synergistic additive and novel properties for sensor development. This paper covers more than 290 research works since 2015, elaborating the diverse roles played by various nanomaterials in the biosensing field. Hence, we provide a comprehensive review of the healthcare sensing applications of nanomaterials, covering carbon allotrope-based, inorganic, and organic nanomaterials. These sensing systems are able to detect a wide variety of clinically relevant molecules, like nucleic acids, viruses, bacteria, cancer antigens, pharmaceuticals and narcotic drugs, toxins, contaminants, as well as entire cells in various sensing media, ranging from buffers to more complex environments such as urine, blood or sputum. Thus, the latest advancements reviewed in this paper hold tremendous potential for the application of nanomaterials in the early screening of diseases and point-of-care testing

Development and Application of a Predictive Model to Detect Analyte Concentration from a Quartz Crystal Microbalance

This work characterizes a sensor technology employing thin coating film deposited on a Quartz Crystal Microbalance (QCM) device. The specific objective of this technology is to predict analyte concentration in rapid response. This work develops ion exchange/adsorption models to interpret sensor signal to analyte concentration. Several practical cases from industry or literature were studied. Emphasis has been placed on chemical and biochemical analyte detection. This work has demonstrated that the proposed method can rapidly predict analyte concentration with continuous step-changes, which will significantly strengthen the QCM sensor technology.School of Chemical Engineerin

Bioengineering and biomechanical approaches for pancreatic cancer

Pancreatic cancer is the fourth-leading cause of cancer related mortality and is predicted to be the second leading cause of cancer death by 2030. A hallmark feature of pancreatic ductal adenocarcinoma (PDAC) is dense fibrotic stroma surrounding the tumor, composed of extracellular matrix (ECM) and cells such as myofibroblasts. The properties of this stroma and functional contribution to carcinogenesis and disease progression has been the subject of intense focus in the past decade; yet, the role of mechanobiology in modulating the phenotype of immune cells in the tumor microenvironment remains to be elucidated. Although a lack of understanding PDAC etiology and progression limits effective treatments that can be deployed by clinicians, current methods of diagnosing PDAC likely are insufficient even if such treatments exist, especially if there is a narrow early window for drug efficacy. Recently, however, extracellular vesicles have emerged as powerful circulating blood biomarkers, thus paving the way for a new era of non-invasive cancer diagnostics. However, currently the process of extracellular vesicle isolation and detection is not only highly inefficient, but also technically challenging. This thesis describes bioengineering tools and biomechanical investigations of pancreatic cancer. In Chapter 2, the biomechanical phenotype of macrophages is studied in context of a stromal modulation agent, the chemotherapeutic drug tamoxifen. Tamoxifen was found to regulate macrophage focal adhesion dynamics, cytoskeletal activity, migratory behavior, and expression of TLR4. In Chapter 3, a novel microfluidic device was modeled and built to determine cell adhesion strength with potential applications to investigate regulation of focal adhesion structure by candidate drugs. Chapter 4 describes the development of methods and devices for isolation and detection of extracellular vesicles using acoustophoresis and a graphene field effect transistor, respectively. Such tools and perspectives could serve to detect PDAC earlier as well as identify and test new therapies.Open Acces

Electrochemical biosensor based on microfabricated electrode arrays for life sciences applications

In developing a biosensor, the utmost important aspects that need to be emphasized are the specificity and selectivity of the transducer. These two vital prerequisites are of paramount in ensuring a robust and reliable biosensor. Improvements in electrochemical sensors can be achieved by using microelectrodes and to modify the electrode surface (using chemical or biological recognition layers to improve the sensitivity and selectivity). The fabrication and characterisations of silicon-based and glass-based gold microelectrode arrays with various geometries (band and disc) and dimension (ranging from 10 μm-100 nm) were reported. It was found that silicon-based transducers of 10 μm gold microelectrode array exhibited the most stable and reproducible electrochemical measurements hence this dimension was selected for further study. Chemical electrodeposition on both 10 μm microband and microdisc were found viable by electro-assisted self-assembled sol-gel silica film and nanoporous-gold electrodeposition respectively. The fabrication and characterisations of on-chip electrochemical cell was also reported with a fixed diameter/width dimension and interspacing variation. With this regard, the 10 μm microelectrode array with interspacing distance of 100 μm exhibited the best electrochemical response. Surface functionalisations on single chip of planar gold macroelectrodes were also studied for the immobilisation of histidine-tagged protein and antibody. Imaging techniques such as atomic force microscopy, fluorescent microscopy or scanning electron microscope were employed to complement the electrochemical characterisations. The long-chain thiol of self-assembled monolayer with NTA-metal ligand coordination was selected for the histidine-tagged protein while silanisation technique was selected for the antibody immobilisation. The final part of the thesis described the development of a T-2 labelless immunosensor using impedimetric approach. Good antibody calibration curve was obtained for both 10 μm microband and 10 μm microdisc array. For the establishment of the T-2/HT-2 toxin calibration curve, it was found that larger microdisc array dimension was required to produce better calibration curve. The calibration curves established in buffer solution show that the microelectrode arrays were sensitive and able to detect levels of T-2/HT-2 toxin as low as 25 ppb (25 μg kg-1) with a limit of quantitation of 4.89 ppb for a 10 μm microband array and 1.53 ppb for the 40 μm microdisc array

Minimally invasive diagnosis of Alzheimer’s disease by detecting microRNA using a quartz crystal resonator

In 2014, there were 850,000 people living with dementia in the UK, creating an economic burden of £26.3 billion a year. 62% of dementia patients are diagnosed with Alzheimer’s Disease (AD). AD is a slow progressing disease with three phases: a long prodromal stage followed by mild cognitive impairment and then late AD. The prodromal phase of AD is on average 30 years. So, when the first symptoms become apparent, the pathology in the brain is extensive. If Alzheimer’s could be diagnosed early, when the pathological load is lower, this may improve the chances of finding a disease modifying therapy.For diagnosis during the prodromal stage to be viable, patients would need to be diagnosed through mass screening, with the most appropriate diagnosis method being biomarker detection in peripheral blood. There are an increasing number of articles in literature researching the use of non coding RNA sequences called microRNA (miRNA) as biomarkers for AD. However, their viability in diagnosing prodromal AD is unknown and the current miRNA detection method, the polymerase chain reaction (PCR), is time consuming, expensive and requires experienced personnel, making it unsuitable for use in mass screening. Therefore, the aim of the thesis was to investigate miRNA as a prodromal biomarker with a new detection method to determine the usability of miRNA as a prodromal AD biomarker.AD is characterised by the build up of amyloid β and hyper phosphorylated tau in the brain. The movement of tau through the brain is divided into 6 Braak stages. Chapter 4 aimed to determine the point at which miRNAs deregulate in AD. To achieve this, post mortem brain tissue was obtained through all 6 Braak stages. The RNA from the post-mortem brain samples were isolated and the change in miRNA levels were determined using PCR and the ΔΔct method. After comparing miRNA to a spike in control the 4 miRNAs tested showed no significant change in miRNA levels through the progression of AD.One of the first signs of AD is the activation of astrocytes in the brain. The aim of Chapter 5 was to determine the effect of the deregulated miRNAs on astrocytes. An astrocytoma cell line was activated using lipopolysaccharides and TNF α then transfected so specific miRNA were over expressed. The concentration of metabolites, cytokines and growth factors were measured in the cell supernatant. Results showed mir 210 regulated G CSF and mir 223 regulated glutamate consumption.Finally, the feasibility of rapid detection of miRNA was investigated using a quartz crystal microblance (QCM). The QCM, assembled within a custom built microfluidic flow cell, was driven at its fundamental resonance frequency. Shifts in the third Fourier harmonic current and the resonance frequency were measured for a range of concentrations of a single stranded DNA (ssDNA). The results show feasibility for rapid quantitative detection of ssDNA to 60 ng/mL, in an easy to use label free assay. Amplification of the signal was seen with the addition of electrochemical potential and the use of particles.Results looking at the deregulation of miRNA in the temporal cortex showed no significant change, therefore further work is needed looking at alternative miRNA sequences. The results with ssDNA suggest that quartz crystal resonator has the potential for rapid, specific and quantitative miRNA detection. However, amplification strategies would help to achieve the clinically relevant limit of detection in peripheral blood.</div

Probing multivalent particle–surface interactions using a quartz crystal resonator

The rise in market-approved cellular therapies demands for advancements in process analytical technology (PAT) capable of fulfilling the requirements of this new industry. Unlike conventional biopharmaceuticals, cell-based therapies (CBT) are complex “live” products, with a high degree of inherent biological variability. This exacerbates the need for in-process monitoring and control of critical product attributes, in order to guarantee safety, efficacious and continuous supply of this CBT. There are therefore mutual industrial and regulatory motivations for high throughput, non-invasive and non-destructive sensors, amenable to integration in an enclosed automated cell culture system. While a plethora of analytical methods is available for direct characterization of cellular parameters, only a few satisfy the requirements for online quality monitoring of industrial-scale bioprocesses. [Continues.