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Switchable control of protein adhesion on silicon oxide surfaces
Frank Meiners, , University of Oldenburg
The control of cell and protein adhesion plays an important role in many areas like biomedical devices or biosensors. In order to create surfaces that do not allow for cell adhesion, coatings of oligoethylene glycol (OEG) and poly(ethylene glycol) (PEG) units are used. To prevent the cell adhesion on silicon surfaces this surfaces can be modified by using OEG- and PEG-terminated alkylsilanes. One important property is that these OEG- and PEG-terminated surfaces can be switched on to cell and protein adhesion by oxidation.
In this work, the modifications of silicon oxide with the methyl-capped OEG-alkyltrichlorosilane are described. After the modification it is possible to switch on the protein adhesion by using UV/ozone treatment.
Development of a Microfluidic Device for Efficient Protein Extraction
Eman Alzahrani, Student, University Of Hull
a new process for the fabrication of a silica-based monolith inside a glass microchip and its modification with octadecylsilyl ligands was successfully developed for use in the microchip-based solid phase extraction of proteins. Monolithic porous silica without cracks was prepared by a modified sol-gel process, followed by placement of the silica monolith disk inside the extraction chamber in the base plate of the microchip. The two plates of the glass microchip were then thermally bonded at 575 °C for 3 hours. The silica monolith was not affected by the thermal bonding of the two plates of the microchip. This process completely avoids the problem of shrinkage in the silica skeleton during preparation. The monolithic silica inside the glass microchip was subsequently modified with octadecylsilyl (C18) moieties for increased protein binding capacity. The performance of the microchip was evaluated using the extraction of six proteins varying in molecular weight and isoelectric point, namely insulin, cytochrome C, lysozyme, myoglobin, ß-lactoglobulin, and hemoglobin at a concentration of 60 µM. The results show that the octadecylated silica monolith was permeable, has the ability to remove impurities, and achieved a high extraction recovery of the proteins (94.8 - 99.7%) compared with conventional octadecylated silica particles (48.3 - 91.3%).
High-throughput optical injection of mammalian cells using 2-D hydrodynamic focusing and a non-diffracting beam
Helen Rendall, Phd Student, University of St Andrews
Femtosecond photoporation is an optical method for the injection of membrane impermeable substances into cells. Typically this is a low-throughput method where each cell is individually targeted. We build on previous work which utilises a microfluidic platform for automated optical injection. In this new geometry, 2-dimensional hydrodynamic focusing is achieved using a 3-dimensional nozzle which confines mammalian cells to the central region of the microfluidic channel. A reusable quartz chip is designed so that a Bessel beam can be directed along the centre of the channel, parallel to the flow of cells. This allows for higher flow speeds to be used whilst maintaining an adequate dwell time in the core of the non-diffracting beam compared to the previous, orthogonal approach. Using this method we have achieved viable injection of HL60 cells with propidium iodide with an efficiency of 20±4% at a rate of 10 cells/sec where several thousand cells are collected within 15 minutes.
Microdevices for molecular diagnostics
Johannes Peham, Postdoc, Austrian Institute of Technology
Miniaturised devices based on molecular methods have the great potential to improve diagnostic tests and biomarker discovery in terms of rapidness, throughput, sensitivity and cost-efficiency. We present a tubing-based continuous-flow PCR system for the high-throughput analysis of long DNA targets up to 1.3 kbp. Sensitivities down to 100 cells per reaction were achieved and throughputs of 80 samples per hour could be realised. Fluorescent labelling of nucleotides was performed in a disposable lab on a chip device. The disc-shaped system amplifies and labels DNA in a single reaction, accelerating the workflow of DNA microarray analysis. The device is disposable, enabled by injection moulding and thermal bonding, which are technologies capable for large-scale production. An overall process acceleration from 6 h to 1.5 h could be achieved with the lab on a chip system at sensitivites down to 100 bacterial cells per reaction. The two systems are superior to current assays in terms of sample throughput, rapidness, process-integration and cost-efficiency. This enables improved biomarker identification and advanced diagnostic assays.
Rails and anchors: guiding and trapping drops in 2D using wells of surface energy
Rémi Dangla, Chief Executive Officer, Ecole Polytechnique
We present a method to control nanolitre drops confined in a wide and thin microchannels, by etching fine patterns into the channel’s top surface. It relies on the fact that the droplets reduce their surface energy as they enter into a local depression. The resulting gain in free energy pulls them into the groove. We show experimentally that localized holes can be used as anchors for holding drops, while linear patterns can be used as rails to guide them along complex trajectories.
The anchor
Optical Detection of Glucose Concentrations in a PDMS Enzymatic Biosensor
Alaaldeen Al-Halhouli, Research Associate, Technische Universität Braunschweig
Microsystem technology for life science applications has become increasingly important in medicine and biotechnology. Such systems include a micro bioreactor (MBR) for cultivation, microfluidic devices for fluid handling and a biosensor for detecting product concentrations. The general challenge of such systems is the lack of reliable data on reactant and product concentrations across the downstream of the MBRs.
This paper presents a PDMS enzymatic biosensor that consists of a microbeads chamber, beads filling channel, reactor chamber, self-alignment structures for the fiber optics with micro-lenses. Since the glucose concentrations can be optically measured as a function of absorbed light, TMB mediator is used. The mediator changes its colour to green due to the reduction of H2O2. These proportional changes indicate on the glucose concentration. Experimentally, several glucose concentrations were continuously pumped at a 33 µL/h and measurements were reported after 10 and 15 min. Results showed that the sensor is able to detect glucose concentrations up to 10 mM with a standard deviation of 0.96 - 0.98 and a limit of detection of 0.7 mM. Such results are very promising for the application in optical LOC systems used for online analysis or optic MBRs.
A novel “Clamp and Play” platform for automated biological protocols
Maïwenn Kersaudy-Kerhoas, Research Associate, Heriot-Watt University
The automation of biological and chemical protocols necessitate the development of tools to avoid human errors, cross-contamination and lengthy procedures. Requirements for these tools also include versatility, cost, and accessibility. Microfluidic devices which present these features are highly desirable solutions.
In this work, we present an original “clamp and play” platform featuring pneumatic, heating and magnetic controls acting on a physically detachable microfluidic chip. This automated microfluidic platform has the unique advantage of eliminating the need for tubing on the chip and is fully compatible with high throughput multichannel pipettes. Based on this decoupling concept, a disposable plastic microfluidic chip can be simply clamped onto the permanent platform containing a pneumatic fluid actuation system, heating and magnetic controls for the full automation of biological protocols.
Performance testing on the flow rates, magnetic and temperature controls demonstrates the capability of our platform to perform high precision fluid operations for biological protocol automation in a low cost, contamination-free mode.
Towards miniaturised tools for non-invasive prenatal monitoring
Maïwenn Kersaudy-Kerhoas, Research Associate, Heriot-Watt University
Traditionally, methods to sample foetal material for diagnosis are amniocentesis and chorionic villus (CVS). The material obtained is analyzed for cytogenetic, molecular and biochemical abnormalities [1]. Both procedures are invasive, with associated risks to the mother and foetus. Amniocentesis leads to miscarriage in approximately 1% of cases and CVS in around 1–2% of cases [2-3].
Miniaturised integrated solutions are presented permitting non-invasive prenatal diagnosis (NIPD) from maternal blood sample. A microfluidic system allowing the extraction of plasma from maternal blood using only passive structures was validated for NIPD use in a clinical study. Quantitative polymerase chain reaction was performed on SRY gene primers in the on-chip extracted maternal plasma to differentiate male and female foetuses. Blind results were compared to the gender examinations at birth.
Independent on-chip DNA amplification was conducted in stationary and continuous flow based devices for potential validation in a complete NIPD system. The SRY gene was successfully amplified within a stationary flow using rapidly prototyped PMMA microPCR device. A standard 30 PCR cycles were completed within 30 minutes and amplification products were confirmed by agarose gel based electrophoresis.
Monolithic integrated photonic system
Angie Ma, Candidate, University College London
We report an innovative microsystem in which waveguides arrays, fibre-to-waveguide couplers and microfluidic channels are integrated to form a complete microchip for photonic characterisation of micron-sized objects. The waveguides arrays allows high resolution optical measurements. In addition to imaging, the scattering of an opaque object(a silicon cantilever tip) in the microfluidic channel was studied. The diffraction pattern obtained through the waveguides contains information about the object's physical properties, including size and position to a high degree of sensitivity. So far we are able to resolve objects of size down to 3um and position change of 0.5um. We anticipate smaller objects can be resolved especially if we move towards a shorter wavelength. To our knowledge, this is the first time the diffraction pattern within the size regime of microfluidic systems was measured using integrated optics. Applications to this technique are broad including the device can be adapted for studying biological samples such as cells and proteins. In general, the device offers the possibility of eliminating external free space optical components which are often required with lab on chip devices.
EIS cell chip to monitor cell behaviour and migration
Elisabetta Primiceri, , CNR Istituto Nanoscienze/University of Salento
An important goal of biomedical research is the development of tools for high-throughput evaluation of drug effects and cytotoxicity. In this respect, electrochemical impedance spectroscopy (EIS) is an emerging technique for fabrication of new biosensors and lab-on-chips. One of the most interesting application concerns the study of cells since RET and C correlated with viability, adhesion and cytoskeleton with the great advantage of real-time monitoring without any damage for cells.
We have developed devices for different applications such as cell counting, viability tests and aptotactic assays: in particular the last one deals with automatic migration assays. This biochip, inspired by a Boyden chamber, consists of two compartments separated by a porous membrane and it is aligned to EIS sensors. Cells, seeded in the upper chamber through a microfluidic channel, can migrate through the pores of the membrane to the lower chamber and get in touch with the electrodes to be detected. As proof of principle hepatocellular carcinoma (HCC) cells have been studied as a function of microenvironment. The results reveal that the cell chip provides an easy and real-time monitoring tool to study cellular processes such as migration, and to perform cell counting and viability tests.
Impedentiometric Protein Chip for Label Free Immunosensor applications
Maria Serena Chiriaco, , University of Salento
Impedance biosensors based on the immobilization of molecules on electrodes surface could be used as a powerful tool to investigate interactions between biomolecules. This could be a new skill for diagnostic purposes and could be used in clinical analysis and POC tests because of its low cost and label free features. Biosensors based on Electrochemical Impedance Spectroscopy (EIS) technique take advantage on the possibility to modulate the capacitance C and interfacial electron transfer resistance RET, measured at the electrodes surface in correlation to the interaction between molecules. Biorecognition events as for example the binding of an antigen on its specific antibody could be then monitored and studied [1]. Based on a strong experience on biosensors [2-4] we developed a protein chip able to detect both antigens or antibodies thanks to a different functionalization of electrodes on the basis of the species researched. In particular we realized a cholera toxin immunosensor able to detect the toxin from acqueous solutions [5] and a protein chip for the easy detection of antigens from serum samples. A microfluidic system completes the setup of our device with the presence of channels and enclosed reaction chambers, and make it a good platform for on chip analysis.
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