Portable dielectrophoresis for biology: ADEPT facilitates cell trapping, separation, and interactions.

Dielectrophoresis is a powerful and well-established technique that allows label-free, non-invasive manipulation of cells and particles by leveraging their electrical properties. The practical implementation of the associated electronics and user interface in a biology laboratory, however, requires an engineering background, thus hindering the broader adoption of the technique. In order to address these challenges and to bridge the gap between biologists and the engineering skills required for the implementation of DEP platforms, we report here a custom-built, compact, universal electronic platform termed ADEPT (adaptable dielectrophoresis embedded platform tool) for use with a simple microfluidic chip containing six microelectrodes. The versatility of the open-source platform is ensured by a custom-developed graphical user interface that permits simple reconfiguration of the control signals to address a wide-range of specific applications: (i) precision positioning of the single bacterium/cell/particle in the micrometer range; (ii) viability-based separation by achieving a 94% efficiency in separating live and dead yeast; (iii) phenotype-based separation by achieving a 96% efficiency in separating yeast and Bacillus subtilis; (iv) cell-cell interactions by steering a phagocytosis process where a granulocyte engulfs E. coli RGB-S bacterium. Together, the set of experiments and the platform form a complete basis for a wide range of possible applications addressing various biological questions exploiting the plug-and-play design and the intuitive GUI of ADEPT.

Broadband stripline Lenz lens achieves 11 × NMR signal enhancement.

A Lenz lens is an electrically passive conductive element that, when placed in a time-varying magnetic field, acts as a magnetic flux concentrator or a magnetic lens. In the realm of nuclear magnetic resonance (NMR), Lenz lenses have been exploited as electrically passive metallic radiofrequency interposers placed between a sample and a tuned or untuned NMR detector in order to focus the [Formula: see text]-field of the detector onto a smaller sample space. Here we explore a novel embodiment of the Lenz lens, which acts as a non-resonant stripline interposer, i.e., the [Formula: see text]-field acts along the longitudinal volume of a sample container, such as a capillary or other microfluidic channel that is coincident with the axis of the stripline. The almost vanishing self-resonance of the stripline Lenz lens, at frequencies relevant for NMR, leads to a desirable [Formula: see text]-field amplitude that is nearly perfectly uniform across the sample and hence lacking a characteristic sinusoidal modal shape. The action of Lenz' law ensures that no stray [Formula: see text]-field is found outside of the stripline's active volume. Because the stripline Lenz lens does not rely on its own geometry to achieve resonance, its frequency response is thus widely broadband for field enhancements up to a factor of 11, with only the external driving resonator properties governing the overall resonant behaviour. We explore the use of the stripline Lenz lens with a sub-nanolitre sample volume, readily detecting 4 isotopes with resonances ranging from 125.76 to 500 MHz. The concept holds potential for the NMR study of thin films, small biological samples, as well as the in situ study of battery materials.

Surface characterisation reveals substrate suitability for cyanobacterial phototaxis.

Cyanobacteria respond to light stimulation, activating localised assembly of type IV pili for motility. The resulting phototactic response is highly dependent on the nature of the incoming light stimulus, and the final motility parameters depend on the surface properties. Conventionally, phototaxis studies are carried out on hydrogel surfaces, such as agarose, with surface properties that vary in time due to experimental conditions. This study considers five substrates, widely utilized in microfluidic technology, to identify the most suitable alternative for performing reliable and repeatable phototaxis assays. The surfaces are characterised via a contact angle goniometer to determine the surface energy, white light interferometry for roughness, zeta-potentials and AFM force distance curves for charge patterns, and XPS for surface composition. Cell motility assays showed 1.25 times increment on surfaces with a water contact angle of 80° compared to a reference glass surface. To prove that motility can be enhanced, polydimethylsiloxane (PDMS) surfaces were plasma treated to alter their surface wettability. The motility on the plasma-treated PDMS showed similar performance as for glass surfaces. In contrast, untreated PDMS surfaces displayed close to zero motility. We also describe the force interactions of cells with the test surfaces using DLVO (Derjaguin-Landau-Verwey-Overbeek) and XDLVO (extended DLVO) theories. The computed DLVO/XDLVO force-distance curves are compared with those obtained using atomic force microscopy. Our findings show that twitching motility on tested surfaces can be described mainly from adhesive forces and hydrophobicity/hydrophilicity surface properties. STATEMENT OF SIGNIFICANCE: The current article focuses on unravelling the potential Micro-Electro-Mechanical System (MEMS) compatible surfaces for studying phototactic twitching motility of cyanobacteria. This is the first exhaustive surface characterization study coupled with phototaxis experiments, to understand the forces contributing to twitching motility. The methods shown in this paper can be further extended to study other surfaces and also to other bacteria exhibiting twitching motility.

Real-Time NMR Monitoring of Spatially Segregated Enzymatic Reactions in Multilayered Hydrogel Assemblies*.

Compartmentalized chemical reactions at the microscale are important in biotechnology, yet monitoring the molecular content at these small scales is challenging. To address this challenge, we integrate a compact, reconfigurable reaction cell featuring electrochemical functionality with high-resolution NMR spectroscopy. We demonstrate the operation of this system by monitoring the activity of enzymes immobilized in chemically distinct layers within a multi-layered chitosan hydrogel assembly. As a benchmark, we observed the parallel activities of urease (Urs), catalase (Cat), and glucose oxidase (GOx) by monitoring reagent and product concentrations in real-time. Simultaneous monitoring of an independent enzymatic process (Urs) together with a cooperative process (GOx + Cat) was achieved, with chemical conversion modulation of the GOx + Cat process demonstrated by varying the order in which the hydrogel was assembled.

Untuned broadband spiral micro-coils achieve sensitive multi-nuclear NMR TX/RX from microfluidic samples.

The low frequency plateau in the frequency response of an untuned micro-resonator permits broadband radio-frequency reception, albeit at the expense of optimal signal-to-noise ratio for a particular nucleus. In this contribution we determine useful figures of merit for broadband micro-coils, and thereby explore the parametric design space towards acceptable simultaneous excitation and reception of a microfluidic sample over a wide frequency band ranging from 13C to 1H, i.e., 125-500 MHz in an 11.74 T magnet. The detector achieves 37% of the performance of a comparably sized, tuned and matched resonator, and a linewidth of 17 ppb using standard magnet shims. The use of broadband detectors circumvents numerous difficulties introduced by multi-resonant RF detector circuits, including sample loading effects on matching, channel isolation, and field distortion.

An NMR-compatible microfluidic platform enabling in situ electrochemistry.

Combining microfluidic devices with nuclear magnetic resonance (NMR) has the potential of unlocking their vast sample handling and processing operation space for use with the powerful analytics provided by NMR. One particularly challenging class of integrated functional elements from the perspective of NMR are conductive structures. Metallic electrodes could be used for electrochemical sample interaction for example, yet they can cause severe NMR spectral and SNR degradation. These issues are more entangled at the micro-scale since the distorted volume occupies a higher ratio of the sample volume. In this study, a combination of simulation and experimental validation was used to identify an electrode geometry that, in terms of NMR spectral parameters, performs as well as for the case when no electrodes are present. By placing the metal tracks in the side-walls of a microfluidic channel, we found that NMR RF excitation performance was actually enhanced, without compromising B0 homogeneity. Monitoring in situ deposition of chitosan in the microfluidic platform is presented as a proof-of-concept demonstration of NMR characterisation of an electrochemical process.

Inductively coupled magic angle spinning microresonators benchmarked for high-resolution single embryo metabolomic profiling.

The magic angle coil spinning (MACS) technique has been introduced as a very promising extension for solid state NMR detection, demonstrating sensitivity enhancements by a factor of 14 from the very first time it has been reported. The main beneficiary of this technique is the scientific community dealing with mass- and volume-limited, rare, or expensive samples. However, more than a decade after the first report on MACS, there is a very limited number of groups who have continued to develop the technique, let alone it being widely adopted by practitioners. This might be due to several drawbacks associated with the MACS technology until now, including spectral linewidth, heating due to eddy currents, and imprecise manufacturing. Here, we report a device overcoming all these remaining issues, therefore achieving: (1) spectral resolution of approx 0.01 ppm and normalized limit of detection of approx. 13 nmol s0.5 calculated using the anomeric proton of sucrose at 3 kHz MAS frequency; (2) limited temperature increase inside the MACS insert of only 5 °C at 5 kHz MAS frequency in an 11.74 T magnetic field, rendering MACS suitable to study live biological samples. The wafer-scale fabrication process yields MACS inserts with reproducible properties, readily available to be used on a large scale in bio-chemistry labs. To illustrate the potential of these devices for metabolomic studies, we further report on: (3) ultra-fine 1H-1H and 13C-13C J-couplings resolved within 10 min for a 340 mM uniformly 13C-labeled glucose sample; and (4) single zebrafish embryo measurements through 1H-1H COSY within 4.5 h, opening the gate for the single embryo NMR studies.

Spatial scanning hyperspectral imaging combining a rotating slit with a Dove prism.

We present a new concept for spatial scanning hyperspectral imaging. Spatial scanning is one of the main methods used for hyperspectral data acquisition and can provide high spectral resolution over a wide spectral range. However, conventional techniques, such as the whiskbroom and the pushbroom techniques, suffer from the need for relative motion between the target and the imaging system, which increases the complexity on the hardware side and limits the application possibilities. Our new approach combines a rotating slit and a co-rotating Dove prism. The rotating slit scans the target image by selecting one line from the image at each angular position of the slit. The rotating Dove prism is used to synchronously re-align the transmitted light from the selected image line with respect to the transmission grating to allow the projection of the diffracted light over the same range of pixel columns of the image sensor to facilitate data acquisition and extraction of spectral information. The new approach enables the spatial scanning of the target image without the need for relative linear motion or the use of additional external equipment and therefore opens the door for more application scenarios.

Broadband and multi-resonant sensors for NMR.

It has always been of considerable interest to study the nuclear magnetic resonance response of multiple nuclei simultaneously, whether these signals arise from internuclear couplings within the same molecule, or from uncoupled nuclei within sample mixtures. The literature contains numerous uncorrelated reports on techniques employed to achieve multi-nuclear NMR detection. This paper consolidates the subset of techniques in which single coil detectors are utilized, and highlights the strengths and weaknesses of each approach, at the same time pointing the way towards future developments in the field of multi-nuclear NMR. We compare the different multi-nuclear NMR techniques in terms of performance, and present a guide to NMR probe designers towards application-based optimum design. We also review the applicability of micro-coils in the context of multi-nuclear methods. Micro-coils benefit from compact geometries and exhibit lower impedance, which provide new opportunities and challenges for the NMR probe designer.

"Small is beautiful" in NMR.

In this prospective paper we consider the opportunities and challenges of miniaturized nuclear magnetic resonance. As the title suggests, (irreverently borrowing from E.F. Schumacher's famous book), miniaturized NMR will feature a few small windows of opportunity for the analyst. We look at what these are, speculate on some open opportunities, but also comment on the challenges to progress.

Spatial and Temporal Control Over Multilayer Bio-Polymer Film Assembly and Composition.

Biosensing applications have taken advantage of lab-on-a-chip technologies for sample handling and sensors integration for highly sensitive, specific detection with high throughput. These systems are based on 2.5D fabrication principles with sensing elements restricted to an array format in two dimensions. In this report, a sensing platform that recovers biosensing capabilites in three spatial dimensions is presented. This is achieved by leveraging chitosan, a stimulus responsive polyaminosaccharide that undergoes a sol-gel transition driven by a change of pH. This process can be repeated, resulting in a multilayered hydrogel stack where each layer carries a unique chemical identity. In addition, the functionality of chitosan can be modified prior to or during the assembly process. This is demonstrated by introducing both a carboxylic acid functionality and additional primary amines to the base chitosan polymer. The assembly process is shown to be compatible with microfluidic dimensions.

A Non-Resonant Kinetic Energy Harvester for Bioimplantable Applications.

A linear non-resonant kinetic energy harvester for implantable devices is presented. The design contains a metal platform with permanent magnets, two stators with three-dimensional helical coils for increased power generation, ball bearings, and a polydimethylsiloxane (PDMS) package for biocompatibility. Mechanical excitation of this device within the body due to daily activities leads to a relative motion between the platform and stators, resulting in electromagnetic induction. Initial prototypes without packaging have been fabricated and characterized on a linear shaker. Dynamic tests showed that the friction force acting on the platform is on the order of 0.6 mN. The resistance and the inductance of the coils were measured to be 2.2 Ω and 0.4 µH, respectively. A peak open circuit voltage of 1.05 mV was generated per stator at a platform speed of 5.8 cm/s. Further development of this device offers potential for recharging the batteries of implantable biomedical devices within the body.

Dual-mode pushbroom hyperspectral imaging using active system components and feed-forward compensation.

Among the methods developed for hyperspectral imaging, pushbroom spatial scanning stands out when it comes to achieving high spectral resolution over a wide spectral range. However, conventional pushbroom systems are usually realized using passive system components, which has limited their flexibility and adaptability and narrowed their application scenarios. In this work, we adopt a different approach to the design and construction of pushbroom systems based on using active internal components. We present a new system concept utilizing an internal line scanning unit and a rotating camera mechanism. This enables a dual-mode imaging system that allows switching between 2D spatial imaging and spectral imaging. The line scanning unit, which consists of a narrow slit mounted to a linear piezo motor, facilitates the spatial scanning of the target while eliminating the laborious relative motion between the target and the imaging system, which is needed in conventional spectrographs. A software is developed for the automation and synchronization of the active components, which enables a novel feed-forward compensation function to compensate the shift in the diffraction angle due to the scanning motion of the slit and provide higher flexibility in data acquisition.

Heteronuclear Micro-Helmholtz Coil Facilitates µm-Range Spatial and Sub-Hz Spectral Resolution NMR of nL-Volume Samples on Customisable Microfluidic Chips.

We present a completely revised generation of a modular micro-NMR detector, featuring an active sample volume of ∼ 100 nL, and an improvement of 87% in probe efficiency. The detector is capable of rapidly screening different samples using exchangeable, application-specific, MEMS-fabricated, microfluidic sample containers. In contrast to our previous design, the sample holder chips can be simply sealed with adhesive tape, with excellent adhesion due to the smooth surfaces surrounding the fluidic ports, and so withstand pressures of ∼2.5 bar, while simultaneously enabling high spectral resolution up to 0.62 Hz for H2O, due to its optimised geometry. We have additionally reworked the coil design and fabrication processes, replacing liquid photoresists by dry film stock, whose final thickness does not depend on accurate volume dispensing or precise levelling during curing. We further introduced mechanical alignment structures to avoid time-intensive optical alignment of the chip stacks during assembly, while we exchanged the laser-cut, PMMA spacers by diced glass spacers, which are not susceptible to melting during cutting. Doing so led to an overall simplification of the entire fabrication chain, while simultaneously increasing the yield, due to an improved uniformity of thickness of the individual layers, and in addition, due to more accurate vertical positioning of the wirebonded coils, now delimited by a post base plateau. We demonstrate the capability of the design by acquiring a 1H spectrum of ∼ 11 nmol sucrose dissolved in D2O, where we achieved a linewidth of 1.25 Hz for the TSP reference peak. Chemical shift imaging experiments were further recorded from voxel volumes of only ∼ 1.5 nL, which corresponded to amounts of just 1.5 nmol per voxel for a 1 M concentration. To extend the micro-detector to other nuclei of interest, we have implemented a trap circuit, enabling heteronuclear spectroscopy, demonstrated by two 1H/13C 2D HSQC experiments.

Microfabricated inserts for magic angle coil spinning (MACS) wireless NMR spectroscopy.

This article describes the development and testing of the first automatically microfabricated probes to be used in conjunction with the magic angle coil spinning (MACS) NMR technique. NMR spectroscopy is a versatile technique for a large range of applications, but its intrinsically low sensitivity poses significant difficulties in analyzing mass- and volume-limited samples. The combination of microfabrication technology and MACS addresses several well-known NMR issues in a concerted manner for the first time: (i) reproducible wafer-scale fabrication of the first-in-kind on-chip LC microresonator for inductive coupling of the NMR signal and reliable exploitation of MACS capabilities; (ii) improving the sensitivity and the spectral resolution by simultaneous spinning the detection microcoil together with the sample at the "magic angle" of 54.74° with respect to the direction of the magnetic field (magic angle spinning - MAS), accompanied by the wireless signal transmission between the microcoil and the primary circuit of the NMR spectrometer; (iii) given the high spinning rates (tens of kHz) involved in the MAS methodology, the microfabricated inserts exhibit a clear kinematic advantage over their previously demonstrated counterparts due to the inherent capability to produce small radius cylindrical geometries, thus tremendously reducing the mechanical stress and tearing forces on the sample. In order to demonstrate the versatility of the microfabrication technology, we have designed MACS probes for various Larmor frequencies (194, 500 and 700 MHz) testing several samples such as water, Drosophila pupae, adamantane solid and LiCl at different magic angle spinning speeds.

Lab on a chip phased-array MR multi-platform analysis system.

We present a lab on a chip (LOC) compatible modular platform for magnetic resonance (MR)-based investigation of sub-millimetre samples. The platform combines the advantages offered respectively by microcoils (high resolution at the microscale) and macroscopic surface coils (large field of view) as MR-detectors and consists of a phased array of microcoils (PAMs) providing a flat MR-sensitive area of 18.3 mm(2) with a B(0)-field uniformity better than 0.25 ppm in the sensor centre area. We demonstrate both high-resolution magnetic resonance imaging (MRI) and NMR spectroscopy using this platform. To demonstrate the application for biological samples, we report MR imaging of fish oocytes with an in-plane resolution of 30 × 30 µm(2) and a contrast to noise ratio of 10 for a scan time of only 13 min 39 s. We have also demonstrated high-resolution spectroscopy of a water phantom achieving 11 ppb (4.5 Hz at 400 MHz) linewidth and an SNR of 28 for only 12 s scan time. State of the art automatic wire bonding technology in conjunction with MEMS techniques has been employed to manufacture the platform with potential applications in MR-investigation of planar samples.

Complex three-dimensional high aspect ratio microfluidic network manufactured in combined PerMX dry-resist and SU-8 technology.

In this paper we present a new fabrication method that combines for the first time popular SU-8 technology and PerMX dry-photoresist lamination for the manufacturing of high aspect ratio three-dimensional multi-level microfluidic networks. The potential of this approach, which further benefits from wafer-level manufacturing and accurate alignment of fluidic levels, is demonstrated by a highly integrated three-level microfluidic chip. The hereby achieved network complexity, including 24 fluidic vias and 16 crossing points of three individual microchannels on less than 13 mm(2) chip area, is unique for SU-8 based fluidic networks. We further report on excellent process compatibility between SU-8 and PerMX dry-photoresist which results in high interlayer adhesion strength. The tight pressure sealing of a fluidic channel (0.5 MPa for 1 h) is demonstrated for 150 µm narrow SU-8/PerMX bonding interfaces.

On-chip three dimensional microcoils for MRI at the microscale.

We present for the first time a fully MEMS-integrated technology to manufacture 3D geometrically perfect solenoidal microcoils for microscale MRI applications. We report 25 microm isotropic resolution MR images of a copper sulfate aqueous phantom. These images are acquired using microcoils with 5 windings of insulated 25 microm diameter Au wire and with quality factors as high as 46 at the operating frequency (400 MHz).