Membrane-based microfluidic systems for medical and biological applications.

Microfluidic devices with integrated membranes that enable control of mass transport in constrained environments have shown considerable growth over the last decade. Membranes are a key component in several industrial processes such as chemical, pharmaceutical, biotechnological, food, and metallurgy separation processes as well as waste management applications, allowing for modular and compact systems. Moreover, the miniaturization of a process through microfluidic devices leads to process intensification together with reagents, waste and cost reduction, and energy and space savings. The combination of membrane technology and microfluidic devices allows therefore magnification of their respective advantages, providing more valuable solutions not only for industrial processes but also for reproducing biological processes. This review focuses on membrane-based microfluidic devices for biomedical science with an emphasis on microfluidic artificial organs and organs-on-chip. We provide the basic concepts of membrane technology and the laws governing mass transport. The role of the membrane in biomedical microfluidic devices, along with the required properties, available materials, and current challenges are summarized. We believe that the present review may be a starting point and a resource for researchers who aim to replicate a biological phenomenon on-chip by applying membrane technology, for moving forward the biomedical applications.

Corrigendum: Challenge of material haemocompatibility for microfluidic blood-contacting applications.

[This corrects the article DOI: 10.3389/fbioe.2023.1249753.].

Challenge of material haemocompatibility for microfluidic blood-contacting applications.

Biological applications of microfluidics technology is beginning to expand beyond the original focus of diagnostics, analytics and organ-on-chip devices. There is a growing interest in the development of microfluidic devices for therapeutic treatments, such as extra-corporeal haemodialysis and oxygenation. However, the great potential in this area comes with great challenges. Haemocompatibility of materials has long been a concern for blood-contacting medical devices, and microfluidic devices are no exception. The small channel size, high surface area to volume ratio and dynamic conditions integral to microchannels contribute to the blood-material interactions. This review will begin by describing features of microfluidic technology with a focus on blood-contacting applications. Material haemocompatibility will be discussed in the context of interactions with blood components, from the initial absorption of plasma proteins to the activation of cells and factors, and the contribution of these interactions to the coagulation cascade and thrombogenesis. Reference will be made to the testing requirements for medical devices in contact with blood, set out by International Standards in ISO 10993-4. Finally, we will review the techniques for improving microfluidic channel haemocompatibility through material surface modifications-including bioactive and biopassive coatings-and future directions.

Low-Cost, Accessible Fabrication Methods for Microfluidics Research in Low-Resource Settings.

Microfluidics are expected to revolutionize the healthcare industry especially in developing countries since it would bring portable, easy-to-use, self-contained diagnostic devices to places with limited access to healthcare. To date, however, microfluidics has not yet been able to live up to these expectations. One non-negligible factor can be attributed to inaccessible prototyping methods for researchers in low-resource settings who are unable to afford expensive equipment and/or obtain critical reagents and, therefore, unable to engage and contribute to microfluidics research. In this paper, we present methods to create microfluidic devices that reduce initial costs from hundreds of thousands of dollars to about $6000 by using readily accessible consumables and inexpensive equipment. By including the scientific community most embedded and aware of the requirements of healthcare in developing countries, microfluidics will be able to increase its reach in the research community and be better informed to provide relevant solutions to global healthcare challenges.

Thermoplastic elastomer with advanced hydrophilization and bonding performances for rapid (30 s) and easy molding of microfluidic devices.

One of the most important areas of research on microfluidic technologies focuses on the identification and characterisation of novel materials with enhanced properties and versatility. Here we present a fast, easy and inexpensive microstructuration method for the fabrication of novel, flexible, transparent and biocompatible microfluidic devices. Using a simple hot press, we demonstrate the rapid (30 s) production of various microfluidic prototypes embossed in a commercially available soft thermoplastic elastomer (sTPE). This styrenic block copolymer (BCP) material is as flexible as PDMS and as thermoformable as classical thermoplastics. It exhibits high fidelity of replication using SU-8 and epoxy master molds in a highly convenient low-isobar (0.4 bar) and iso-thermal process. Microfluidic devices can then be easily sealed using either a simple hot plate or even a room-temperature assembly, allowing them to sustain liquid pressures of 2 and 0.6 bar, respectively. The excellent sorption and biocompatibility properties of the microchips were validated via a standard rhodamine dye assay as well as a sensitive yeast cell-based assay. The morphology and composition of the surface area after plasma treatment for hydrophilization purposes are stable and show constant and homogenous distribution of block nanodomains (∼22° after 4 days). These domains, which are evenly distributed on the nanoscale, therefore account for the uniform and convenient surface of a "microfluidic scale device". To our knowledge, this is the first thermoplastic elastomer material that can be used for fast and reliable fabrication and assembly of microdevices while maintaining a high and stable hydrophilicity.

Heavy social drinkers score higher on implicit wanting and liking for alcohol than alcohol-dependent patients and light social drinkers.

BACKGROUND AND OBJECTIVES: Automatic hedonic ("liking") and incentive ("wanting") processes are assumed to play an important role in addiction. Whereas some neurobiological theories suggest that these processes become dissociated when drug use develops into an addiction (i.e., "liking" becomes weaker, whereas "wanting" becomes exaggerated; e.g., Robinson & Berridge, 1993), other theories suggest that there is a linear relationship between these two processes (i.e., both "liking" and "wanting" increase equally; e.g., Koob & Le Moal, 1997). Our aim was to examine "wanting" and "liking" in three groups of participants: alcohol-dependent patients, heavy social drinkers, and light social drinkers. METHODS: Participants performed two different single target implicit association tests (ST-IATs; e.g., Bluemke & Friese, 2007) and explicit ratings that were designed to measure "liking" and "wanting" for alcohol. RESULTS: Our results are in sharp contrast with the theories of both Robinson and Berridge and Koob and Le Moal: heavy drinkers had higher scores than light drinkers and alcohol-dependent patients on both the wanting ST-IAT and the liking ST-IAT. There were no differences between alcohol-dependent patients and light drinkers. Explicit ratings mirrored these results. LIMITATIONS: These findings suggest that our ST-IATs are not valid measures of "wanting" and "liking". Instead, they might assess more complex knowledge regarding participants' experiences and goals. CONCLUSIONS: These findings suggest that the relationship between drug consumption and appetitive drug associations is not linear, highlighting the importance of testing both sub-clinical and clinical samples in future research.

From cellular lysis to microarray detection, an integrated thermoplastic elastomer (TPE) point of care Lab on a Disc.

We present an all-thermoplastic integrated sample-to-answer centrifugal microfluidic Lab-on-Disc system (LoD) for nucleic acid analysis. The proposed CD system and engineered platform were employed for analysis of Bacillus atrophaeus subsp. globigii spores. The complete assay comprised cellular lysis, polymerase chain reaction (PCR) amplification, amplicon digestion, and microarray hybridization on a plastic support. The fluidic robustness and operating efficiency of the assay were ensured through analytical optimization of microfluidic tools enabling beneficial implementation of capillary valves and accurate control of all flow timing procedures. The assay reliability was further improved through the development of two novel microfluidic strategies for reagents mixing and flow delay on the CD platform. In order to bridge the gap between the proof-of-concept LoD and production prototype demonstration, low-cost thermoplastic elastomer (TPE) was selected as the material for CD fabrication and assembly, allowing the use of both, high quality hot-embossing and injection molding processes. Additionally, the low-temperature and pressure-free assembly and bonding properties of TPE material offer a pertinent solution for simple and efficient loading and storage of reagents and other on-board components. This feature was demonstrated through integration and conditioning of microbeads, magnetic discs, dried DNA buffer reagents and spotted DNA array inserts. Furthermore, all microfluidic functions and plastic parts were designed according to the current injection mold-making knowledge for industrialization purposes. Therefore, the current work highlights a seamless strategy that promotes a feasible path for the transfer from prototype toward realistic industrialization. This work aims to establish the full potential for TPE-based centrifugal system as a mainstream microfluidic diagnostic platform for clinical diagnosis, water and food safety, and other molecular diagnostic applications.

Dissociation of cardiogenic and postnatal myocardial activities of GATA4.

Transcription factor GATA4 is a critical regulator of the embryonic and postnatal heart, but the mechanisms and cofactors required for its diverse functions are not fully understood. Here, we show that whereas the N-terminal domain of GATA4 is required for inducing cardiogenesis and for promoting postnatal cardiomyocyte survival, distinct residues and domains therein are necessary to mediate these effects. Cardiogenic activity of GATA4 requires a 24-amino-acid (aa) region (aa 129 to 152) which is needed for transcriptional synergy and physical interaction with BAF60c. The same region is not essential for induction of endoderm or blood cell markers by GATA4, suggesting that it acts as a cell-type-specific transcriptional activation domain. On the other hand, a serine residue at position 105, which is a known target for mitogen-activated protein kinase (MAPK) phosphorylation, is necessary for GATA4-dependent cardiac myocyte survival and hypertrophy but is entirely dispensable for GATA4-induced cardiogenesis. We find that S105 is differentially required for transcriptional synergy between GATA4 and serum response factor (SRF) but not other cardiac cofactors such as TBX5 and NKX2.5. The findings provide new insight into GATA4 mechanisms of action and suggest that distinct regulatory pathways regulate activities of GATA4 in embryonic development and postnatal hearts.

3D thermoplastic elastomer microfluidic devices for biological probe immobilization.

Microfluidics has emerged as a valuable tool for the high-resolution patterning of biological probes on solid supports. Yet, its widespread adoption as a universal biological immobilization tool is still limited by several technical challenges, particularly for the patterning of isolated spots using three-dimensional (3D) channel networks. A key limitation arises from the difficulties to adapt the techniques and materials typically used in prototyping to low-cost mass-production. In this paper, we present the fabrication of thin thermoplastic elastomer membranes with microscopic through-holes using a hot-embossing process that is compatible with high-throughput manufacturing. The membranes provide the basis for the fabrication of highly integrated 3D microfluidic devices with a footprint of only 1 × 1 cm(2). When placed on a solid support, the device allows for the immobilization of up to 96 different probes in the form of a 10 × 10 array comprising isolated spots of 50 × 50 µm(2). The design of the channel network is optimized using 3D simulations based on the Lattice-Boltzmann method to promote capillary action as the sole force distributing the liquid in the device. Finally, we demonstrate the patterning of DNA and protein arrays on hard thermoplastic substrates yielding spots of excellent definition that prove to be highly specific in subsequent hybridization experiments.

Thermoplastic elastomers for microfluidics: towards a high-throughput fabrication method of multilayered microfluidic devices.

Multilayer soft lithography of polydimethylsiloxane (PDMS) is a well-known method for the fabrication of complex fluidic functions. With advantages and drawbacks, this technique allows fabrication of valves, pumps and micro-mixers. However, the process is inadequate for industrial applications. Here, we report a rapid prototyping technique for the fabrication of multilayer microfluidic devices, using a different and promising class of polymers. Using styrenic thermoplastic elastomers (TPE), we demonstrate a rapid technique for the fabrication and assembly of pneumatically driven valves in a multilayer microfluidic device made completely from thermoplastics. This material solution is transparent, biocompatible and as flexible as PDMS, and has high throughput thermoforming processing characteristics. We established a proof of principle for valving and mixing with three different grades of TPE using an SU-8 master mold. Specific viscoelastic properties of each grade allow us to report enhanced bonding capabilities from room temperature bonding to free pressure thermally assisted bonding. In terms of microfabrication, beyond classically embossing means, we demonstrate a high-throughput thermoforming method, where TPE molding experiments have been carried out without applied pressure and vacuum assistance within an overall cycle time of 180 s. The quality of the obtained thermoplastic systems show robust behavior and an opening/closing frequency of 5 Hz.

Rapid isothermal substrate microfabrication of a biocompatible thermoplastic elastomer for cellular contact guidance.

The use of microstructured substrates to study and influence cell orientation, which plays an important role in tissue functionality, has been of great interest lately. Silicon and poly(dimethylsiloxane) substrates have typically been used, but long processing times and exogenous protein surface coating, required to enhance cell viability, limit their use as large-scale platforms. There is thus a need for a non-biodegradable biocompatible substrate that allows rapid and low cost microfabrication. In this paper a styrene-(ethylene/butylene)-styrene block co-polymer (SEBS) microstructured by a rapid replication technique using low pressure an isothermal hot embossing approach has been demonstrated. SEBS substrates were treated with oxygen plasma to enhance cell adhesion and sterilized using ethylene oxide gas. While cell adhesion to and proliferation on these substrates was as good as on tissue culture polystyrene, cellular alignment on microstructured SEBS was also very high (97.7±0.5%) when calculated within a 10° angle variation from the longitudinal axis. Furthermore, tissue sheets on microstructured SEBS have been produced and exhibited cellular alignment within the engineered tissue. In addition, these results were obtained without coating the material with exogenous proteins. Such substrates should be helpful in the culture of tissue engineered substitutes with an intrinsic orientation and to elucidate questions in cell biology.

Surface topography induces 3D self-orientation of cells and extracellular matrix resulting in improved tissue function.

The organization of cells and extracellular matrix (ECM) in native tissues plays a crucial role in their functionality. However, in tissue engineering, cells and ECM are randomly distributed within a scaffold. Thus, the production of engineered-tissue with complex 3D organization remains a challenge. In the present study, we used contact guidance to control the interactions between the material topography, the cells and the ECM for three different tissues, namely vascular media, corneal stroma and dermal tissue. Using a specific surface topography on an elastomeric material, we observed the orientation of a first cell layer along the patterns in the material. Orientation of the first cell layer translates into a physical cue that induces the second cell layer to follow a physiologically consistent orientation mimicking the structure of the native tissue. Furthermore, secreted ECM followed cell orientation in every layer, resulting in an oriented self-assembled tissue sheet. These self-assembled tissue sheets were then used to create 3 different structured engineered-tissue: cornea, vascular media and dermis. We showed that functionality of such structured engineered-tissue was increased when compared to the same non-structured tissue. Dermal tissues were used as a negative control in response to surface topography since native dermal fibroblasts are not preferentially oriented in vivo. Non-structured surfaces were also used to produce randomly oriented tissue sheets to evaluate the impact of tissue orientation on functional output. This novel approach for the production of more complex 3D tissues would be useful for clinical purposes and for in vitro physiological tissue model to better understand long standing questions in biology.

An exploration of the moderating effect of trait emotional intelligence on memory and attention in neutral and stressful conditions.

Trait emotional intelligence (trait EI) refers to individual differences in the experience, perception, regulation, and utilization of emotions. Research has shown that trait EI moderated subjective and endocrine responses to both natural and laboratory stressors. This study explores, the cognitive processes underlying this effect, under the hypothesis that trait EI moderates the impact of stress on memory and/or attention. Results supported the hypothesis, but solely for the 'regulation' EI-dimension (named self-control or SC). In neutral conditions, high SC was characterized by an attentional focus to neutral material and a facilitated memory for positive events, whereas low SC was characterized by an attentional focus to emotional material (regardless of valence) and a facilitated memory for negative events. In stressful conditions, high SC individuals engaged attention to emotional material (regardless of valence) and recalled more negative events, while low SC individuals disengaged attention from emotional material and recalled more positive events.

Microfluidic patterning of miniaturized DNA arrays on plastic substrates.

This paper describes the patterning of DNA arrays on plastic surfaces using an elastomeric, two-dimensional microcapillary system (muCS). Fluidic structures were realized through hot-embossing lithography using Versaflex CL30. Like elastomers based on poly(dimethylsiloxane), this thermoplastic block copolymer is able to seal a surface in a reversible manner, making it possible to confine DNA probes with a level of control that is unparalleled using standard microspotting techniques. We focus on muCSs that support arrays comprising up to 2 x 48 spots, each being 45 mum in diameter. Substrates were fabricated from two hard thermoplastic materials, poly(methylmethacrylate) and a polycyclic olefin (e.g., Zeonor 1060R), which were both activated with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide hydrochloride and N-hydroxysuccinimide to mediate covalent attachment of DNA molecules. The approach was exemplified by using 0.25-32 muM solutions of amino-modified oligonucleotides labeled with either Cy3 or Cy5 fluorescent dye in phosphate-buffered saline, allowing for a direct and sensitive characterization of the printed arrays. Solutions were incubated for durations of 1 to >48 h at 22, 30, and 40 degrees C to probe the conditions for obtaining uniform spots of high fluorescence intensity. The length (l) and depth (d) of microfluidic supply channels were both important with respect to depletion as well as evaporation of the solvent. While selective activation of the substrate proved helpful to limit unproductive loss of oligonucleotides along trajectories, incubation of solution in a humid environment was necessary to prevent uncontrolled drying of the liquid, keeping the immobilization process intact over extended periods of time. When combined, these strategies effectively promoted the formation of high-quality DNA arrays, making it possible to arrange multiple probes in parallel with a high degree of uniformity. Moreover, we show that resultant arrays are compatible with standard hybridization protocols, which allowed for reliable discrimination of individual strands when exposed to a specific ssDNA target molecule.

Relationship between alexithymia, alexithymia factors and salivary cortisol in men exposed to a social stress test.

BACKGROUND: The fact that alexithymia is associated with several medical and psychiatric disorders suggests that it may be a vulnerability factor for various diseases, possibly by enhancing stress responses. To test this "alexithymia-stress hypothesis", we measured the influence of alexithymia and alexithymia subfactors on the cortisol response to an acute stressor. METHODS: Twenty-eight male students were exposed to the Trier Social Stress Test (TSST), during which saliva samples for cortisol determination were collected. RESULTS: Subjects reacted to the stressor with a significant cortisol response. Subjects scoring high on alexithymia evidenced an increased basal anticipatory cortisol level but their peak cortisol and area under the curve were similar to that of low scorers. Multiple regression analyses revealed that the increased cortisol in high scorers was due to only one subfactor of alexithymia, "the difficulty in describing feelings" factor (DDF). DDF high scorers reacted with a large increase in cortisol during anticipation but not during exposure to the stress test. CONCLUSION: The observation that alexithymia scores were associated with differences in cortisol levels before social stress exposure raises the possibility that alexithymia modulates cortisol levels, possibly by affecting the anticipatory cognitive appraisal of situations. This may be essentially attributed to the DDF factor. This observation sheds new light on the "alexithymia-stress hypothesis", which may be of importance to better understand the relationship between alexithymia and diseases. Further studies to address this issue should focus on the factorial structure of the construct and on the importance of anticipation.

Resilience and hypothalamic-pituitary-adrenal axis reactivity under acute stress in young men.

The present study examined the relationship between resilience (measured using the Resilience Scale for Adults) and hypothalamic-pituitary-adrenal (HPA) axis reactivity. We examined the subjective and cortisol responses of 28 healthy young men to an acute stressor (public speech task). Eight saliva samples were collected in order to obtain the response curve (anticipation, reactivity, recuperation) for each subject. ANOVA indicated that highly resilient individuals tended to display less mood deterioration than less resilient individuals (marginal p(time x group interaction) = 0.075). They also revealed that the former tended to secrete less cortisol overall than the latter during the experiment (marginal p(main group effect) = 0.087) but this effect was not uniform across time (p(time x group interaction) = 0.029). Additional analyses performed to identify the source of this interaction revealed that resilience moderates cortisol secretion in anticipation of the stressor (i.e. highly resilient individuals secreted less cortisol than less resilient ones, p = 0.05) but that it is not conductive to lower HPA reactivity amidst stress (i.e. there was no difference between groups in the increase in cortisol secretion from baseline to peak). The recovery slopes were likewise not statistically different. The implications of these findings regarding health are discussed.

The moderating impact of emotional intelligence on free cortisol responses to stress.

The construct of trait emotional intelligence (trait EI) refers to the individual differences in the perception, processing, regulation and utilization of emotional information. Several studies have found that trait EI was a significant moderator of subjective responses (e.g., mood deterioration, emotional intensity, action tendencies, bodily sensations) to both natural and laboratory stressors. The present study aims at extending these findings by examining whether trait EI also moderates the biological (i.e., cortisol) response to stress. To this end, 56 participants were assigned to either a neutral or a stressful condition (public speech task) and psychological and cortisol reactivity were examined. Results revealed that higher trait EI scores were associated with significantly lower reactivity to stress at both psychological (i.e., mood deterioration) and biological (i.e., salivary cortisol) levels. Additional analyses revealed that trait EI had incremental validity to predict stress reactivity over and above social desirability, alexithymia and the five-factor model of personality.

Microfluidic ELISA on non-passivated PDMS chip using magnetic bead transfer inside dual networks of channels.

Achieving efficient passivation of micro-channels against non-specific adsorption of biomolecules is a critical aspect in the development of microfluidic ELISA systems. Usual surface treatments such as pre-coating of the channels with serum albumin, exposure to oxygen plasma, polyethylene glycol grafting however exhibit a lack of long-term stability, with procedures that can be time-consuming, complex or associated with costly materials and instruments. In this paper, we present a new fluidic design combined with an original strategy of manipulating magnetic beads in order to reduce assay noise in bead-based microfluidic ELISA without the need for prior channel pre-treatment. The novelty of the system relies on the physical separation of the immune complex formation phase and the enzymatic reaction phase into two independent networks of channels. These networks are linked by fluidic bridges, whose openings are controlled by pressure valves, and through which the beads are magnetically transferred. A standard curve for the quantification of a model antibody was obtained within 30 minutes. A detection limit of 100 pg mL(-1) (660 fM) and good linearity of the signal up to 4 ng mL(-1) were observed.

Surface modification of thermoplastics--towards the plastic biochip for high throughput screening devices.

Microarrays have become one of the most convenient tools for high throughput screening, supporting major advances in genomics and proteomics. Other important applications can be found in medical diagnostics, detection of biothreats, drug discovery, etc. Integration of microarrays with microfluidic devices can be highly advantageous in terms of portability, shorter analysis time and lower consumption of expensive biological analytes. Since fabrication of microfluidic devices using traditional materials such as glass is rather expensive, there is great interest in employing polymeric materials as a low cost alternative that is suitable for mass production. A number of commercially available plastic materials were reviewed for this purpose and poly(methylmethacrylate) Zeonor 1060R and Zeonex E48R were identified as promising candidates, for which methods for surface modification and covalent immobilization of DNA oligonucleotides were developed. In addition, we present proof-of-concept plastic-based microarrays with and without integration with microfluidics.