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The development of new immunotherapeutic drugs and combinatorial strategies requires the implementation of novel methods to test their efficacy in vitro. Here, we present a series of miniaturized in vitro assays to assess immune cell cytotoxic activity, infiltration, and phenotype in renal carcinoma spheroids with the use of a recently developed multichambered microwell chip. We provide protocols for tumor spheroid formation, NK cell culture, fluorescence labelling and imaging of live or fixed cells directly in the chip together with data analysis., Part of ISBN 9780443191633QC 20231116

Examen i vårdadministration, YH-utbildning: 20 YH-poäng

Here, we present a methodology based on multiplexed fluorescence screening of two-or three-dimensional cell cultures in a newly designed multichambered microwell chip, allowing direct assessment of drug or im-mune cell cytotoxic efficacy. We establish a framework for cell culture, formation of tumor spheroids, fluores-cence labeling, and imaging of fixed or live cells at various magnifications directly in the chip together with data analysis and interpretation. The methodology is demonstrated by drug cytotoxicity screening using ovarian and non-small cell lung cancer cells and by cellular cytotoxicity screening targeting tumor spheroids of renal carcinoma and ovarian carcinoma with natural killer cells from healthy donors. The miniaturized format allowing long-term cell culture, efficient screening, and high-quality imaging of small sample volumes makes this methodology promising for individualized cytotoxicity tests for precision medicine., QC 20230215

Miniaturization of cell culture substrates enables controlled analysis of living cells in confined micro-scale environments. This is particularly suitable for imaging individual cells over time, as they can be monitored without escaping the imaging field-of-view (FoV). Glass materials are ideal for most microscopy applications. However, with current methods used in life sciences, glass microfabrication is limited in terms of either freedom of design, quality, or throughput. In this work, we introduce laser-induced deep etching (LIDE) as a method for producing glass microwell arrays for live single cell imaging assays. We demonstrate novel microwell arrays with deep, high-aspect ratio wells that have rounded, dimpled or flat bottom profiles in either single-layer or double-layer glass chips. The microwells are evaluated for microscopy-based analysis of long-term cell culture, clonal expansion, laterally organized cell seeding, subcellular mechanics during migration and immune cell cytotoxicity assays of both adherent and suspension cells. It is shown that all types of microwells can support viable cell cultures and imaging with single cell resolution, and we highlight specific benefits of each microwell design for different applications. We believe that high-quality glass microwell arrays enabled by LIDE provide a great option for high-content and high-resolution imaging-based live cell assays with a broad range of potential applications within life sciences., QC 20230220

Despite advancements in analytical technologies, their complexity and cost have largely restricted their application in scalable online or multiplexed measurements. Here we report a quartz crystal resonator (QCR)-based method for detection of macromolecules that allows immensely simpler and faster measurements by employing for the first time a fixed frequency drive (FFD) and analytical expressions of acoustic parameters. Using human immunoglobulin E (hIgE) as an exemplar macromolecule and an anti-hIgE aptamer functionalised on a QCR, quantitative accuracy was benchmarked against the traditional impedance analysis method. The ability of FFD to capture data over longer observation periods at significantly higher acquisition rates at a fixed amplitude showed improvement in the QCR's sensitivity and specificity of transduction. The foundations for low-cost and low power online integration and large-scale multiplexability are also discussed., QC 20220429

Glass materials have excellent optical and chemical properties for microscopy-based live cell assays but state-of-the-art methods for microfabrication of Lab-on-Chip (LoC) devices are often limited by either complex manufacturing and/or low quality results. In this work, we have evaluated glass microwell array chips produced using a recently introduced laser-based microfabrication method. Three different types of microwell designs have been tested for imaging and screening of on-chip cell cultures and live cell assays., Part of ISBN 9781733419031QC 20230614

Direct detection of whole vegetative bacteria was investigated employing a quartz crystal resonator (QCR) in its nonlinear regime. Escherichia coli (E. coli) in buffer solution under flow was captured on a QCR in a microfluidic cell using a whole-cell anti-E.coli aptamer. The nonlinear distortion in QCR response due to the 'pull' from surface-bound bacteria was measured in realtime as the change in third Fourier harmonic (3f) current and compared with shifts in the traditional acoustic parameters of resonance frequency and dissipation. The change in 3f current showed superior quantitative correlation with E. coli concentrations (105-108 cfu/mL) and at least an order of magnitude better sensitivity than shifts in the traditional acoustic parameters. Most interestingly, underpinned by the strength of bacteria-QCR pull, the nonlinear acoustic principle demonstrated a unique specificity in transduction, even in a mixed sample with another gram-negative bacteria, that can supplement the specificity of the bioreceptors. An analytical expression was derived to quantitatively relate the competing influence of shifts in dissipation and nonlinearity coefficient of the QCR on the change in 3f current. This study demonstrates the potential for reliable direct readout of bioreceptor-mediated binding of whole vegetative bacteria from complex samples using a nonlinear acoustic resonator., QC 20200608

Small molecule detection is of wide interest in clinical and industrial applications. However, its accessibility is still limited as miniaturisation and system integration is challenged in reliability, costs and complexity. Here we combined a 14.3 MHz quartz crystal resonator (QCR), actuated and analysed using a fixed frequency drive (FFD) method, with a nanomolecular imprinted polymer for label-free, realtime detection of N-hexanoyl-L-homoserine lactone (199 Da), a gram-negative bacterial infection biomarker. The lowest concentration detected (1 mu M) without any optimisation was comparable with that of a BIAcore SPR system, an expensive laboratory gold standard, with significant enhancement in sensitivity and specificity beyond the state-of-the-art QCR. The analytical formula-based FFD method can potentially allow a multiplexed "QCR-on-chip" technology, bringing a paradigm shift in speed, accessibility and affordability of small molecule detection., QC 20200525

Tuberculosis (TB) infects about 25% of the world population and claims more human lives than any other infectious disease. TB is spread by inhalation of aerosols containing viable Mycobacterium tuberculosis expectorated or exhaled by patients with active pulmonary disease. Air-sampling technology could play an important role in TB control by enabling the detection of airborne M. tuberculosis, but tools that are easy to use and scalable in TB hotspots are lacking. We developed an electrostatic air sampler termed the TB Hotspot DetectOR (THOR) and investigated its performance in laboratory aerosol experiments and in a prison hotspot of TB transmission. We show that THOR collects aerosols carrying microspheres, Bacillus globigii spores and M. bovis BCG, concentrating these microparticles onto a collector piece designed for subsequent detection analysis. The unit was also successfully operated in the complex setting of a prison hotspot, enabling detection of a molecular signature for M. tuberculosis in the cough of inmates. Future deployment of this device may lead to a measurable impact on TB case-finding by screening individuals through the aerosols they generate., QC 20211207

Cell therapy manufacturing is limited by lack of online tools capable of realtime in-process monitoring, particularly of simultaneous changes in multiple orthogonal (mutually independent) parameters. Here, we studied changes in CD36 expression, number density and size (area) of erythroblasts through different stages of erythropoiesis in vitro using a quartz crystal resonator (QCR), integrated with a microscope, and flow cytometry in parallel. An analytical model was developed extending the Kanazawa-Gordon theory. Based on this model, independent correlations were established between changes in each QCR parameter, dissipation (ΔΓ) and resonance frequency (−Δf0), and CD36 expression (from flow cytometry) and cell area (from microscope). The correlation functions were used to derive an acoustic signature (−ΔΓ/Δf0) of the differentiation process that uniquely mapped the relative changes in CD36 expression and late-stage enucleation-related deviations. A method to quantify relative changes in cell area purely from the acoustic parameters was also proposed. This work demonstrated for the first time the potential of an electromechanical tool for online monitoring of concurrently varying orthogonal phenotypic parameters in cell therapy manufacturing. © 2020 The Authors, QC 20210907

NK cells eliminate virus-infected and tumor cells by releasing cytotoxic granules containing granzyme B (GrzB) or by engaging death receptors that initiate caspase cascades. The orchestrated interplay between both cell death pathways remains poorly defined. Here we simultaneously measure the activities of GrzB and caspase-8 in tumor cells upon contact with human NK cells. We observed that NK cells switch from inducing a fast GrzB-mediated cell death in their first killing events to a slow death receptor–mediated killing during subsequent tumor cell encounters. Target cell contact reduced intracellular GrzB and perforin and increased surface-CD95L in NK cells over time, showing how the switch in cytotoxicity pathways is controlled. Without perforin, NK cells were unable to perform GrzB-mediated serial killing and only killed once via death receptors. In contrast, the absence of CD95 on tumor targets did not impair GrzB-mediated serial killing. This demonstrates that GrzB and death receptor–mediated cytotoxicity are differentially regulated during NK cell serial killing., QC 20190822

QC 20211013

A new method for determination of resonance frequency and dissipation of a mechanical oscillator is presented. Analytical expressions derived using the Butterworth-Van Dyke equivalent electrical circuit allow the determination of resonance frequency and dissipation directly from each impedance datapoint acquired at a fixed amplitude and frequency of drive, with no need for numerical fitting or measurement dead time unlike the conventional impedance or ring-down analysis methods. This enables an ultrahigh time resolution and superior noise performance with relatively simple instrumentation. Quantitative validations were carried out successfully against the impedance analysis method for inertial and viscous loading experiments on a 14.3 MHz quartz crystal resonator (QCR). Resonance frequency shifts associated with the transient processes of quick needle touches on a thiol self-assembled-monolayer functionalised QCR in liquid were measured with a time resolution of 112 μs, which is nearly two orders of magnitude better than the fastest reported quartz crystal microbalance. This simple and fast fixed frequency drive (FFD) based method for determination of resonance frequency and dissipation is potentially more easily multiplexable and implementable on a single silicon chip delivering economies of scale., QC 20181129

QC 20181003, Norosensor

Introduction: Quartz crystal microbalance (QCM) is commonly used to study biomolecular binding by measuring shifts in resonance frequency of a quartz-crystal-oscillator. However, the currently used methods like impedance analysis or QCM-D, which require repeated sweeps or ringing, are limited in time resolution (~1 second) due to the need for averaging. This restricts our ability to study transient biomolecular processes, which occur in sub-millisecond time scale. A novel technique has been reported here that allows quantification of resonance frequency of a quartz-crystal-oscillator with significantly improved time resolution by driving and measuring continuously at a constant frequency within the resonance bandwidth. Method: The reactive component of the experimentally obtained impedance is utilized for the estimation of resonance frequency from the Butterworth Van-dyke (BVD) model of a quartz-crystal-oscillator, assuming that changes in motional inductance and capacitance around resonance are negligible. Triplicate sets of experiments involving the binding of streptavidin with a biotin functionalized 14.3 MHz quartz oscillator surface were performed. Intermittent frequency sweeps and fixed frequency drives, both of 0.1 second duration and around 14.3 MHz, were taken at intervals of 2 minutes under the flow of phosphate-buffer-saline (PBS buffer) before and after injection of streptavidin. Results: The average shift in resonance frequency from the baseline (measurements before streptavidin injection) due to streptavidin-biotin binding, calculated from the fixed frequency drive or FFD (148 Hz) was within 1% of that estimated from the frequency sweep method by fitting the experimentally recorded impedance employing the BVD model (149 Hz). Discussion: The agreement of the FFD with conventional frequency sweep method suggests that protein binding can be quantified with reasonable accuracy from each impedance data point, which with our set-up is recorded at 30 kHz s, QC 20190522

Quartz crystal microbalance (QCM) sensing is an established technique commonly used in laboratory based life-science applications. However, the relatively complex, multi-part design and multi-step fabrication and assembly of state-of-the-art QCM cartridges makes them unsuited for disposable applications such as point-of-care (PoC) diagnostics. In this work, we present the uncomplicated manufacturing of QCMs in polymer microfluidic cartridges. Our novel approach comprises two key innovations: the batch reaction injection molding of microfluidic parts; and the integration of the cartridge components by direct, unassisted bonding. We demonstrate molding of batches of 12 off-stoichiometry thiol-ene epoxy polymer (OSTE+) polymer parts in a single molding cycle using an adapted reaction injection molding process; and the direct bonding of the OSTE+ parts to other OSTE+ substrates, to printed circuit boards, and to QCMs. The microfluidic QCM OSTE+ cartridges were successfully evaluated in terms of liquid sealing as well as electrical properties, and the sensor performance characteristics are on par with those of commercially available QCM biosensor cartridge., QC 20171114

The thesis Deathbed Selfie approaches photography and all it entails from a subjective standpoint in an attempt to understand and describe a photographers need to and purpose for photographing. The title of the thesis suggests that all photographs are self-portraits, and eventually they will be autobiographical documents of life as seen by the photographer. After death, images live on as manifestations of a subjective experience. The thesis consists of two parts, a written and an artistic. The written part includes 11 chapters on the various topics that are relevant to the authors personal work. The artistic part includes only a selection of photographs. Through stories based on subjective experiences some of the key issues in photography are illustrated. With these practical examples the author often proves himself wrong, arriving at the conclusion that photography is a complex and everything but black and white medium. Topics that are relevant to contemporary photography are broadly discussed throughout the thesis. These topics include photography as a visual language and the use of narrative in photographic sequences, a photographers moral responsibility, photographs in relation to memory and photographs as proof of a reality. The thesis also discusses the relationship and balance between images and words as a means of communication, and attempts to define the specific act of photographing (taking or making an image). Deathbed Selfie poses the question "why do we photograph?" and seeks to find and answer by studying the photographic process starting from looking at a thing, photographing it and ultimately by looking at the photograph., Tutkielmassa Deathbed Selfie lähestytään valokuvausta ja kaikkea minkä se sisältää subjektiivisesta näkökulmasta pyrkimyksenä kuvailla ja ymmärtää valokuvaajan tarvetta ja syitä valokuvata. Tutkielman nimi vihjaa kaikkien valokuvien olevan omakuvia jotka tulevat lopulta toimimaan omaelämänkerrallisina dokumentaatioina elämästä niin kuin valokuvaaja sen koki. Kuoleman jälkeen valokuvat muutuvat henkilökohtaisen kokemuksen manifestaatioiksi. Työ koostuu yhdestä kirjallisesta ja yhdestä taiteellisesta osuudesta. Kirjallinen osuus sisältää 11 lukua niistä eri aihepiireistä jotka ovat oleellisia kirjoittajan omassa työssä. Taiteellinen osuus koostuu ainoastaan valikoimasta valokuvia. Henkilökohtaisiin kokemuksiin perustuvien tarinoiden avulla havainnollistetaan haastavimmat ongelmat kirjoittajan suhteessa valokuvaukseen. Näiden käytännön esimerkkien avulla kirjoittaja todistaa olevansa itse usein väärässä, todeten että valokuvaus on monimutkainen ja kaikkea paitsi mustavalkoinen taiteenala. Kirjallisessa osuudessa avataan laajasti aiheita jotka liittyy nykyaikaiseen valokuvaukselliseen työhön ja käytäntöihin. Näitä aiheita ovat mm. valokuva visuaalisena kielenä sekä narratiivin hyödyntäminen kuvasarjoissa, valokuvaajan moraalinen vastuu, valokuvan suhde muistiin sekä valokuva todisteena eräästä todellisuudesta. Työ keskustelee myös kuvien ja sanojen suhteesta kommunikaation välineenä sekä pyrkii määrittelemään itse valokuvaamisen tekona. Deathbed Selfie esittää kysymyksen "miksi me valokuvataan?" ja etsii vastausta siihen tutkimalla valokuvauksen prosessia alkaen tietyn asian katsomisesta, sen valokuvaamisesta ja lopulta itse valokuvan tarkastelusta.

In this article, we present OSTE+RIM, a novel reaction injection molding (RIM) process that combines the merits of off-stoichiometric thiol–ene epoxy (OSTE+) thermosetting polymers with the fabrication of high quality microstructured parts. The process relies on the dual polymerization reactions of OSTE+ polymers, where the first curing step is used in OSTE+RIM for molding intermediately polymerized parts with well-defined shapes and reactive surface chemistries. In the facile back-end processing, the replicated parts are directly and covalently bonded and become fully polymerized using the second curing step, generating complete microfluidic devices. To achieve unprecedented rapid processing, high replication fidelity and low residual stress, OSTE+RIM uniquely incorporates temperature stabilization and shrinkage compensation of the OSTE+ polymerization during molding. Two different OSTE+ formulations were characterized and used for the OSTE+RIM fabrication of optically transparent, warp-free and natively hydrophilic microscopy glass slide format microfluidic demonstrator devices, featuring a storage modulus of 2.3 GPa and tolerating pressures of at least 4 bars., QC 201500618

In this work, the development of a point-of-care (PoC) system to capture aerosol from litres of air directly onto a microfluidic lab-on-chip for subsequent analysis is addressed. The system involves an electrostatic precipitator that uses corona charging and electrophoretic transport to capture aerosol droplets onto a microfluidic air-to-liquid interface for downstream analysis. A theoretical study of the governing geometric and operational parameters for optimal electrostatic precipitation is presented. The fabrication of an electrostatic precipitator prototype and its experimental validation using a laboratory-generated aerosolized dye is described. Collection efficiencies were comparable to those of a state-of-the-art Biosampler impinger, with the significant advantage of providing samples that are at least 10 times more concentrated. Finally, we discuss the potential of such a system for breath-based diagnostics., QC 20150223, Rappid

We present a novel integration method for packaging silicon photonic sensors with polymer microfluidics, designed to be suitable for wafer-level production. The method addresses the previously unmet manufacturing challenges of matching the microfluidic footprint area to that of the photonics, and of robust bonding of microfluidic layers to biofunctionalized surfaces. We demonstrate the fabrication, in a single step, of a microfluidic layer in the recently introduced OSTE polymer, and the subsequent unassisted dry bonding of the microfluidic layer to a grating coupled silicon photonic ring resonator sensor chip. The microfluidic layer features photopatterned through holes (vias) for optical fiber probing and fluid connections, as well as molded microchannels and tube connectors, and is manufactured and subsequently bonded to a silicon sensor chip in less than 10 minutes. Combining this new microfluidic packaging method with photonic waveguide surface gratings for light couplin g allows matching the size scale of microfluidics to that of current silicon photonic biosensors., QC 20150928, CellRing, VR-HET, XMEMS

QC 20150401. QC 20160212, Rappid

NQC 2015, Rappid

QC 20150302, Theracage

We present a novel integration method for packaging silicon photonic sensors with polymer microfluidics, designed to be suitable for wafer-level production methods. The method addresses the previously unmet manufacturing challenges of matching the microfluidic footprint area to that of the photonics, and of robust bonding of microfluidic layers to biofunctionalized surfaces. We demonstrate the fabrication, in a single step, of a microfluidic layer in the recently introduced OSTE polymer, and the subsequent unassisted dry bonding of the microfluidic layer to a grating coupled silicon photonic ring resonator sensor chip. The microfluidic layer features photopatterned through holes (vias) for optical fiber probing and fluid connections, as well as molded microchannels and tube connectors, and is manufactured and subsequently bonded to a silicon sensor chip in less than 10 minutes. Combining this new microfluidic packaging method with photonic waveguide surface gratings for light coupling allows matching the size scale of microfluidics to that of current silicon photonic biosensors. To demonstrate the new method, we performed successful refractive index measurements of liquid ethanol and methanol samples, using the fabricated device. The minimum required sample volume for refractive index measurement is below one nanoliter., This paper was published in Optics Express and is made available as an electronic reprint with the permission of OSA. The paper can be found at the following URL on the OSA website:http://dx.doi.org/10.1364/oe.21.021293. Systematic or multiple reproduction or distribution to multiple locations via electronic or other means is prohibited and is subject to penalties under law.Qc 20130913, CellRing

We demonstrate a method for the fast and simple packaging of silicon sensors into a microfluidic package consisting of the recently introduced {OSTE} polymer. The microfluidic layer is first microstructured and thereafter dry-bonded to a silicon photonic sensor, in a process compatible with wafer-level production, and with the entire packaging process lasting only 10 minutes. The fluidic layer combines molded microchannels and fluidic (Luer) connectors with photopatterned through-holes (vias) for optical fiber probing and fluid connections. All the features are fabricated in a single photocuring step. We report measurements with an integrated silicon photonic {Mach-Zehnder} interferometer refractive index sensor packaged by these means., QC 20130718, Cell-Ring

Here we present an uncomplicated and robust method for rapid prototyping of microfluidic devices featuring: 1) lithographically defined, permanently surface modified hydrophilic and hydrophobic regions with contact angles varying from 18o to 118o, in which all four channel walls are surface modified in a single step using polymer chain grafting; 2) polymer chains grafted from open surfaces before bonding, making this method suitable for batch fabrication; 3) biomolecule-compatible, room temperature, dry, homogeneous chip-sealing, in which native as well as hydrophobic/hydrophilic modified OSTE+ surfaces allow for epoxy-epoxy and thiol-epoxy covalent bonding, hence greatly simplifying alignment and dramatically increasing device yields. We demonstrate the method with a functional microfluidic device. This represents a complete, simplified and robust method for batch-manufacturing compatible prototyping of microfluidic devices with tunable mechanical and surface properties., QC 20171122Part of proceedings ISBN: 978-0-9798064-6-9, Theracage

We demonstrate a method for the fast and simple packaging of silicon sensors into a microfluidic package consisting of the recently introduced {OSTE} polymer. The microfluidic layer is first microstructured and thereafter dry-bonded to a silicon photonic sensor, in a process compatible with wafer-level production, and with the entire packaging process lasting only 10 minutes. The fluidic layer combines molded microchannels and fluidic (Luer) connectors with photopatterned through-holes (vias) for optical fiber probing and fluid connections. All the features are fabricated in a single photocuring step. We report measurements with an integrated silicon photonic {Mach-Zehnder} interferometer refractive index sensor packaged by these means., QC 20140212, Cell-Ring

QC 20130521, Rappid

QC 20130521, RPA, Rappid